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Information on EC 4.6.1.18 - pancreatic ribonuclease and Organism(s) Bos taurus and UniProt Accession P61823
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4.6.1.18 pancreatic ribonucleaseIUBMB Comments
Specifically cleaves at the 3'-side of pyrimidine (uracil or cytosine) phosphate bonds in RNA. The reaction takes place in two steps, with the 2',3'-cyclic phosphodiester intermediates released from the enzyme at the completion of the first step. Hydrolysis of these cyclic compounds occurs at a much slower rate through a reversal of the first step, in which the -OH group of water substitutes for the 2'-OH group of the ribose used in the first step, and does not take place until essentially all the susceptible 3',5'-phosphodiester bonds have been cyclised. The enzyme can act as an endo- or exo ribonuclease.
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Word Map
- 4.6.1.18
- asthma
-
allergic
- airway
-
asthmatic
- bronchial
- ige
-
atopic
- pollen
- nasal
- granule
- rhinitis
- sputum
- lavage
-
inhale
-
self-incompatibility
-
expiratory
- eosinophilia
-
degranulation
- histamine
- myeloperoxidase
- tryptase
-
mast
-
leukotriene
-
hyperresponsiveness
-
angiogenin
-
provocation
- methacholine
- pistil
- gametophytic
-
prick
- solanaceae
- rosaceae
-
spirometry
-
nonatopic
-
wheezing
-
ribonucleolytic
-
exhaled
-
fluticasone
- dermatophagoides
-
aeroallergens
-
budesonide
-
nonallergic
-
allergen-induced
- eotaxin
-
interleukin-5
- synthesis
- agriculture
-
hypereosinophilic
- diagnostics
- medicine
-
beclomethasone
-
beta2-agonists
- analysis
- drug development
- pharmacology
-
sil-2r
- s-protein
- biotechnology
The taxonomic range for the selected organisms is: Bos taurus
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea
Reaction Schemes
hide(Overall reactions are displayed. Show all >>)
+
=
+
an [RNA] containing cytidine
+
=
an [RNA]-3'-cytidine-3'-phosphate
+
a 5'-hydroxy-ribonucleotide-3'-[RNA]
+
=
+
an [RNA] containing uridine
+
=
an [RNA]-3'-uridine-3'-phosphate
+
a 5'-hydroxy-ribonucleotide-3'-[RNA]
Synonyms
eosinophil cationic protein, s-rnase, pancreatic ribonuclease, onconase, eosinophil-derived neurotoxin, pancreatic rnase, bovine pancreatic ribonuclease a, bs-rnase, rnase 1, rnase1, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
ribonuclease A
-
RNase A
-
alkaline ribonuclease
-
-
-
-
BS-RNase
Ceratitis capitata alkaline ribonuclease
-
-
-
-
Eosinophil-derived neurotoxin
-
-
-
-
gene S glycoproteins
-
-
-
-
gene S locus-specific glycoproteins
-
-
-
-
glycoproteins, gene S locus-specific
-
-
-
-
glycoproteins, S-genotype-asssocd
-
-
-
-
glycoproteins, SLSG
-
-
-
-
glycoproteins, specific or class, gene S
-
-
-
-
glycoproteins, specific or class, SLSG (gene S locus-specific glycoprotein)
-
-
-
-
nuclease, ribo-
-
-
-
-
pancreatic RNase
ribonuclease
-
-
-
-
ribonuclease A
ribonuclease I
-
-
-
-
Ribonuclease US
-
-
-
-
ribonuclease W1
-
-
-
-
ribonucleate 3'-pyrimidino-oligonucleotidohydrolase
-
-
-
-
ribonucleic phosphatase
-
-
-
-
RL1
-
-
-
-
RNase
-
-
-
-
RNase A
RNase I
-
-
-
-
S-RNase
-
-
-
-
seminal ribonuclease
Seminal RNase
SLSG glycoproteins
-
-
-
-
type I RNase A
-
isoform with the same molecular mass as that of a commercial RNase A
type II RNase A
-
isoform with higher molecular mass than commercial and type I RNase A
pancreatic RNase
-
-
-
-
ribonuclease A
-
-
-
-
ribonuclease A
-
-
RNase A
-
-
-
-
RNase A
-
-
Seminal RNase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
His12, His119 and Lys41 comprise the catalytic site, and several other amino acid residues serve as substrate binding subsites. The mutagenic replacement of Phe120 causes a positional change in His119 and that is a major cause in decreasing the activity of the enzyme. Phe120 is important in fixing the proper spatial position of His119 near the C-terminal region for efficient activity. Phe120 interacts not directly with His119, but with the hydrophobic core
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
the catalytic site of the enzyme is mainly composed of His12, His119, Gln11, Lys41 and Asp121. The conformation of Lys41 and Gln11 are pH dependent and their occurrence is correlated to the release of a sulfate ion from the active site at neutral pH. The sulfate anion is progressively released with pH and water molecules occupy the active site at pH 7.1. The enzyme adopts different conformational states in different pH conditions. In the protonated state, Lys41 points towards the active site both in the presence and in the absence of sulfate and acts as a general base during catalysis
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
the enzyme is composed of 124 amino acid residues. The amino acid residues involved in catalysis are His12, His119 and Lys41. The enzyme has six substrate binding subsites. One of them, B1, binds only to pyrimidine nucleotides and prefers to bind cytidine rather than uridine. Thr45 of the B1 subsite contributes to the enzyme substrate specificity and plays an important role in catalysis. Phe120 in the B1 subsite enhances substrate binding with the aid of its pi electron and hydrophobicity. Asn71 in the B2 subsite is responsible for productive interactions leading to efficient activity, and Glu111 in the same subsite contributes to substrate binding only in the binding of guanosine. Lys7, Arg10 and Gln11 in the P2 subsite also play an important role in catalysis. Some amino acid residues are located in the vicinity of the catalytic residues and contribute to the catalytic reaction by interacting with catalytic residues. Lys7, Arg10 and Lys66 maintain the optimum pKa of His12 and His119. Asp121 positions the proper tautomer of His119. Phe120 is responsible for the strict positioning of His119, and contributes to conformational stability. Tyr97 is involved in maintaining the correct position of Lys41. Cys65-Cys72 are located close to the catalytic and substrate binding residues and contribute to the proper spatial alignment of these residues
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
protein with 124 amino acid residues and four intra-molecular disulfide bonds. In the presence of oxidized and reduced dithiothreitol at pH 8.0 and 25ºC, the enzyme folds through pathways involving a rapid pre-equilibrium resulting in an ensemble of three-disulfide intermediate species. Protein disulfide isomerase catalyzes the conversion of the intermediates to the native enzyme, by acting as both a chaperone and an oxidase on the on-pathway intermediate
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
pyrimidine-specific endoribonuclease which cleaves 3',5'-phosphodiester bonds of single strand RNA via transphosphorylation and subsequent hydrolysis reactions
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
sequential binding of the monomeric substrate in a concentration-dependent manner makes up the active site of the enzyme
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
two isoforms of dimeric enzyme, D-I and D-II
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
His119 and His12 play an important structural role in active site of RNase A
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
enzymatic reaction mechanism, analysis of different versions, e.g. mechanism of Mathias and Rabin for catalysis of the hydrolysis of cytidine 2',3'-cyclic phosphate by RNase A, overview. Model of the RNase A-substrate complex, subsites of RNase A. Bn, Rn, and Rho_n are nucleobase-, ribose-, and phosphate-binding subsites, respectively
-
an [RNA] containing cytidine + H2O = an [RNA]-3'-cytidine-3'-phosphate + a 5'-hydroxy-ribonucleotide-3'-[RNA]
RNase A catalyzes a well-characterized acid-base mechanism involving two histidines (His12 and His119) and a transition state stabilizing positive charge (Lys41). The transphosphorylation of a single-stranded RNA molecule by the enzyme yields a 2',3'-cyclic phosphomonoester intermediate, which can be expelled from the active site or hydrolyzed in a microscopic reverse reaction involving the same two histidines
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hydrolysis of phosphoric ester
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
RNA lyase ([RNA]-3'-cytidine/uridine-3'-phosphate and 5'-hydroxy-ribonucleotide-3'-[RNA] producing)
Specifically cleaves at the 3'-side of pyrimidine (uracil or cytosine) phosphate bonds in RNA. The reaction takes place in two steps, with the 2',3'-cyclic phosphodiester intermediates released from the enzyme at the completion of the first step. Hydrolysis of these cyclic compounds occurs at a much slower rate through a reversal of the first step, in which the -OH group of water substitutes for the 2'-OH group of the ribose used in the first step, and does not take place until essentially all the susceptible 3',5'-phosphodiester bonds have been cyclised. The enzyme can act as an endo- or exo ribonuclease.
CAS REGISTRY NUMBER
COMMENTARY
9001-99-4
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine + H2O
?
-
-
-
?
cyclic 2',3'-nucleoside monophosphate + H2O
3'-phosphomononucleotides
-
second step of hydrolysis is irreversible
24208, 134521, 134522, 134523, 134525, 134527, 134529, 134530, 134531, 134532, 134533, 134534, 134537, 134538, 134541, 134542, 134543, 134551, 134552, 134555, 134563, 134564
-
-
ir
DNA-RNA hybrids + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
in seminal plasma
-
-
?
double-stranded RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
in seminal plasma
-
-
?
poly (C) + H2O
3'-CMP + 3'-phosphooligonucleotides
-
-
-
-
?
poly(A)-poly(U) + H2O
?
-
the enzyme dimers degrade poly(A)-poly(U) dsRNA with an activity that increases with the increase of the oligomer's basicity
-
-
?
RNA + H2O
cyclic 2',3'-nucleoside monophosphate
-
first step of hydrolysis is reversible
24208, 134521, 134522, 134523, 134525, 134527, 134529, 134530, 134531, 134532, 134533, 134534, 134537, 134538, 134541, 134542, 134543, 134551, 134552, 134555, 134563, 134564
-
-
r
cytidine-2',3'-cyclic monophosphate + H2O
3'-CMP
-
hydrolysis reaction
-
-
?
cytidinyl-3',5'-adenosine + H2O
adenosine + 3'-CMP
-
CpA, transphosphorylation reaction
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
endonucleolytic cleavage to 3'-phosphomononucleotides and 3'-phosphooligonucleotides ending in Cp or Up with 2',3'-cyclic phosphate intermediates, e.g. tRNA, 18S and 28S rRNA, yeast RNA,4.5S RNA
24208, 134521, 134522, 134523, 134525, 134527, 134529, 134530, 134531, 134532, 134533, 134534, 134537, 134538, 134541, 134542, 134543, 134551, 134552, 134555, 134556, 134563, 134564
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
high-molecular-weight yeast RNA
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
yeast RNA
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
activity against double-stranded RNA and the antitumour action increase with the size of the oligomer
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
studies on radical scavenging activities of the ribonuclease inhibitor CPRI, a scavenger of pancreatic-type ribonucleases
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
substrate yeast RNA
-
-
?
additional information
?
-
6-carboxyfluorescein-dArU(dA)2-6-carboxytetramethylrhodamine as artificial substrate
-
-
?
additional information
?
-
-
removal of RNA changes the structural stability of low mobility group nonhistone protein (LMG160), in the way that the molecule tends to become self-associated
-
-
?
additional information
?
-
the enzyme degrades single-stranded and/or double-stranded RNA
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
the enzyme degrades single-stranded and/or double-stranded RNA
-
-
?
RNA + H2O
3'-phosphomononucleotides + 3'-phosphooligonucleotides
-
endonucleolytic cleavage to 3'-phosphomononucleotides and 3'-phosphooligonucleotides ending in Cp or Up with 2',3'-cyclic phosphate intermediates, e.g. tRNA, 18S and 28S rRNA, yeast RNA,4.5S RNA
24208, 134521, 134522, 134523, 134525, 134527, 134529, 134530, 134531, 134532, 134533, 134534, 134537, 134538, 134541, 134542, 134543, 134551, 134552, 134555, 134556, 134563, 134564
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Cu2+
Ni2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1-(2,5-dideoxy-5-(4-carboxypiperidinyl)-beta-D-threo-pentofuranosyl)thymine
-
1-(2,5-dideoxy-5-pyrrolidin-1-yl-beta-L-erythro-pentofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione
-
1-(5-deoxy-5-morpholin-4-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
1-(5-deoxy-5-piperidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
1-(5-deoxy-5-pyrrolidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
1-(5-deoxy-5-[4-(ethoxycarbonyl)piperidin-1-yl]-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
2',3'-dideoxy-3'-L-serinylamino thymidine
occupies the active site of ribonuclease A and preferential perturbs the pKa value of His-119 by its free amino group as found from 1H NMR studies, compounds with polar amino acid side chains such as Ser-aT, Tyr-aT and Trp-aT (except His-aT) are more efficient inhibitors compared to those having hydrophobic side chains
2',3'-dideoxy-3'-L-tryptophanylamino thymidine
compounds with polar amino acid side chains such as Ser-aT, Tyr-aT and Trp-aT (except His-aT) are more efficient inhibitors compared to those having hydrophobic side chains
2',3'-dideoxy-3'-L-tyrosylamino thymidine
compounds with polar amino acid side chains such as Ser-aT, Tyr-aT and Trp-aT (except His-aT) are more efficient inhibitors compared to those having hydrophobic side chains
3'-TMP
a competitive inhibitor analogue of the 3'-CMP and 3'-UMP natural product inhibitors, the enzyme shows very high affinty and strong binding with 3'-TMP. Binding of 3'-TMP is very similar to other natural and nonnatural pyrimidine ligands, so single nucleotide affinity is independent of the presence or absence of a 2'-hydroxyl on the ribose moiety of pyrimidines
3-N-piperidine-4-carboxyl-3-deoxy-ara-uridine
binding of two inhibitor molecules in the central cavity of enzyme
4-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-ylcarbamoyl]-butyric acid
-
5'-phospho-2'-deoxyuridine-3-diphosphate (P-5)-adenosine-3'-phosphate
i.e. pdUppA-3'-p, multi-ns molecular dynamics simulations of enzyme in complex with inhibitor
ATP
5-ATP binds with the adenine occupying the B2 subsite in the manner of an RNA substrate but with the gamma-phosphate at the P1 subsite, crystal structure of the complex with pancreatic ribonuclease A
cytosolic ribonuclease inhibitor
RI, from Sus scrofa, binding of the RI molecule to the N-terminal RNase A entity, analysis of crystal structures of the RIRNase A complex and the SGRSGRSG-RNase A tandem enzyme, PDB-ID 1DFJ, overview
-
epicatechin
0.04 mM, 4.4% inhibition, noncompetitve, CD spectral analysis of complex with enzyme, preferred site of binding is around residues 34-39 with possible hydrogen bonding to K7 and R10
epicatechin gallate
0.04 mM, 12.7% inhibition, noncompetitve, CD spectral analysis of complex with enzyme, preferred site of binding is around residues 34-39 with possible hydrogen bonding to K7 and R10
epigallocatechin
0.04 mM, 6.9% inhibition, noncompetitve, CD spectral analysis of complex with enzyme, preferred site of binding is around residues 34-39 with possible hydrogen bonding to K7 and R10
epigallocatechin gallate
0.04 mM, 18.4% inhibition, noncompetitve, CD spectral analysis of complex with enzyme, preferred site of binding is around residues 34-39 with possible hydrogen bonding to K7 and R10
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-malonamic acid
-
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-oxalamic acid
-
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-succinamic acid
-
oligo(vinylsulfonic acid)
potent competitive inhibitor making nearly 8 favorable Coulombic interactions with the enzyme. Oligo(vinylsulfonic acid) is inexpensive and extremely stable. Accoringly oligo(vinylsulfonic acid) has the potential to be useful prophylactic in many chemical, biochemical, and biotechnical experiments involving RNA
P1,P3-bis(5'-adenosyl) triphosphate
crystal structure of the complex with pancreatic ribonuclease A
Pyrophosphate
crystal structure of the complex with pancreatic ribonuclease A
RNasin
for investigating protein translocation in vitro, rough membrane vesicles of endoplasmic reticular origin from the pancreas of different livestock animals can be used as a valuable alternative to the dog source. Since the mRNA in the translation mixture is degraded by ribonucleases present in the membrane fraction, the membrane stocks were diluted in membrane buffer and pretreated with increasing amounts of the recombinant RNase inhibitor RNasin (Promega)
-
thiocyanate
inactivation due to expansion of the enzyme surface and elongation of the catalytic center
3'-CMP
-
strong binding by the wild-type enzyme, reduced binding by enzyme mutants T17A and T82A, kinetics, overview
3-amino-N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-succinamic acid
-
-
aurintricarboxylic acid
-
alters the three-dimensional conformation, dissociation constant of ribonuclease A with aurintricarboxylic acid is 2.33 microM
chitosan
-
molecular weight about 6 kDA, complex formation with enzyme due to establishment of 5-6 ion pairs
diethylpyrocarbonate
-
among the His residues of RNase A, His48 is not accessible to react with diethylpyrocarbonate
Pholiota nameko polysaccharide
-
linear mixed-type inhibition, noncompetitive inhibition is predominant over competitive inhibition
-
ribonuclease inhibitor CPRI
-
scavenger of pancreatic-type ribonucleases, chemiluminescence assay to determine radical scavenging activities toward different reactive oxygen species (ROS) including superoxide anion, hydroxyl radical, lipid-derived radicals and singlet oxygen
-
ribonuclease protein inhibitor
-
the native enzyme is an equilibrium mixture of two isomers, MxM and M=M. In the former the two subunits swap their N-terminal helices. In the reducing environment of the cytosol, isoform M=M dissociates into monomers, which are strongly inhibited by ribonuclease protein inhibitor, wheras isoform MxM remains as a non-covalent dimer which evades ribonuclease protein inhibitor
-
RNase inhibitor
-
RNase A, like most monomeric RNases, is strongly bound and inactivated in mammalian cells by the RNase inhibitor
-
spermidine
-
RNA-binding enzyme activity is regulated through spermidine-induced changes in the charge and structure of the RNA substrate. Spermidine transiently stabilizes RNA sub-populations by binding both specifically and nonspecifically
spermine
-
at 0.13 mM: inhibition, at 0.02 M: activity towards cyclic substrates and poly(C) is activated, not towards poly(U)
Urea
-
mechanism of inhibition, urea inhibits ribonuclease A competitively over a concentration range from 100 mM to 4.0 M, urea with its high dipolar moment is a competitive inhibitor and a very high concentration (more than 4.0 M) of it could denature the enzyme, beginning the interaction with the protein at the active center
cytosolic ribonuclease inhibitor
-
with CpA as substrate, both isoenzymes are fully susceptible to inhibition
-
cytosolic ribonuclease inhibitor
-
protects cells against exogenous ribonucleases, variants of pancreatic ribonuclease that evade ribonuclease inhibitor are cytotoxic, molecular evolution of ribonuclease inhibitor suggests to be a means to enhance the cytotoxicity of mammalian ribonucleases
-
ribonuclease inhibitor
-
extremely tight complex with bovine ribonuclease inhibitor, Kd value 0.69 fM. Kd value of complex with human ribonuclease inhibitor 0.34 fM
ribonuclease inhibitor
-
human placental ribonuclease inhibitor, characterization of its tryptophan residues in the complex with the enzyme. The complex formation results in a more heterogenous environment for both of the optically resolved residues W19 and W375. W19 moves slightly toward a more hydrophobic region, ant the environment of W375 becomes less solvent exposed
additional information
agarose gel and precipitation assays show that the spacer length and the pKa of the carboxylic group have an important role in the inhibitory capacity
-
additional information
enzyme-inhibitor binding and interaction analysis, kinetics, overview
-
additional information
-
enzyme-inhibitor binding and interaction analysis, kinetics, overview
-
additional information
inhibitor synthesis, kinetics, and docking, overview
-
additional information
-
construction of enzyme dimer composed of monomeric units covalently linked by a single amide bond between the side-chains of residues K66 and E9 by incubation of a lyophilized preparation of enzyme under vacuum at 85°C. Dimer exhibits a twofold increase in activity over monomeric enzyme and is not inhibited by the cellular ribonuclease inhibitor protein
-
additional information
-
As(III) species, by avid coordination to the cysteine residues of unfolded reduced proteins, can compromise protein folding pathways, monomethylarsenous acid catalyzes the formation of amyloid-like monodisperse fibrils using reduced ribonuclease A
-
additional information
-
inhibitor synthesis, molecular docking to the enzyme, interaction of inhibitors with hydrogen bonding network formation between His12 and His119 of RNase A, overview
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
sulfate
decreases the distance between the catalytic His residues and increases the globular compactness
alkanediyl-alpha,omega-bis(hydroxyethyl methyl hexadecyl ammonium bromide)
-
the cationic gemini surfactants slightly activate and stabilize RNase A below their critical micelle concentrations at pH 5.0. The cationic gemini surfactant with the shorter spacer interacts more efficiently with RNase A than those with longer spacers, two-transition model, UV, circular dichorism and fluorescence spectroscopies, overview
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.64
cytidine-2',3'-cyclic monophosphate
des-(123-124) mutant, pH 5.5, 25°C
additional information
CpA
-
type I and II isoenzymes, almost identical values to those of commercial enzyme, pH 5.5, 25°C
additional information
cytidine-2',3'-cyclic monophosphate
-
type I and II isoenzymes, almost identical values to those of commercial enzyme, pH 5.5, 25°C
additional information
additional information
-
second-order rate constants for disulfide bond formation in the oxidative folding of RNase A with DHSox as an oxidant at 25°C, overview
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
12
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme M30C/N44C
1044
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme I107C/A122C
1566
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme H105C/V124C
3132
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme A4C/V118C
3480
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme V43C/R85C
4002
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
mutant enzyme R10C/R33C
17400
6-carboxyfluorescein-dArUdAdA-6-carboxytetramethylrhodamine
wild-type enzyme
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.229
1-(2,5-dideoxy-5-(4-carboxypiperidinyl)-beta-D-threo-pentofuranosyl)thymine
pH 7.5, 25°C
0.423
1-(2,5-dideoxy-5-pyrrolidin-1-yl-beta-L-erythro-pentofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione
-
0.179
1-(5-deoxy-5-morpholin-4-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
0.172
1-(5-deoxy-5-piperidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
0.203
1-(5-deoxy-5-pyrrolidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
0.077
1-(5-deoxy-5-[4-(ethoxycarbonyl)piperidin-1-yl]-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione
-
0.25
5'-N-(4-carboxypiperidinyl)-2',3'-didehydro-3',5'-dideoxythymidine
pH 7.5, 25°C
0.38
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-malonamic acid
-
0.132
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-oxalamic acid
-
0.918
N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-succinamic acid
-
0.65
uridine 5'-diphosphate
binds to the active site of the enzyme by anchoring two molecules connected to each other by hydrogen bonds and van der Waals interactions
4
uridine 5'-phosphate
binds to the active site of the enzyme by anchoring two molecules connected to each other by hydrogen bonds and van der Waals interactions
0.037
3-amino-N-[2-hydroxymethyl-5-(5-methyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl)-tetrahydro-furan-3-yl]-succinamic acid
-
pH 7.5, 25°C
0.0039
ribonuclease inhibitor
A4C/C65A/C72A/G88R/V118C mutant, pH 6.0, 25ºC
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
additional information
Bos taurus
-
radical scavenging activities of CPRI indicated, measured by chemiluminescence, values shown, radical scavenging activities higher than those of tea polyphenols
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
1.14
-
substrate poly(A)poly(U), dimethyl suberimidate-treated, monomeric RNase A, pH 7.0, 25°C
13.33
-
substrate yeast RNA, cross-linked RNase A dimer, treated with dimethyl suberimidate, pH 7.0, 25°C
25.21
-
substrate poly(A)poly(U), cross-linked RNase A dimer, treated with dimethyl suberimidate, pH 7.0, 25°C
49.33
-
substrate yeast RNA, dimethyl suberimidate-treated, monomeric RNase A, pH 7.0, 25°C
additional information
additional information
-
isoenzyme D-I shows values of 88 U/mg, 382 U/mg and 6.2 U/mg with yeast RNA, poly(U) and poly(A)poly(U) as substrates, respectively. D-II isoenzyme shows values of 85 U/mg, 387 U/mg and 2.5 U/mg with yeast RNA, poly(U) and poly(A)poly(U) as substrates, respectively
additional information
-
type I isoform, 100% specific activity relative to commercial RNase A. Type II isoform, 53% specific activity relative to commercial enzyme
additional information
-
for assay conditions ribonuclease A solution prepared by dissolving 8 mg ribonuclease A in 4 ml 0.1 M NaHCO3 at pH 9.5 following by pre-cooling to 3°C
additional information
-
enzymatic activities of the various RNase A monomeric and zero-length or domain-swapped oligomeric species on single-stranded yeast or double-stranded poly(A)-poly(U) RNA, overview. The dimers show higher activity compared to the monomners
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
654675, 655979, 656308, 664231, 664308, 664330, 664830, 665486, 666860, 678349, 678519, 678585, 678729, 678745, 678746, 681377, 682742, 682767, 682880, 691341, 706955, 706977, 706984, 707243, 707728, 707730, 707748, 707756, 707757, 709180, 709463, 710510, 710511, 717377, 717596, 718374, 729327, 749474, 749560, 750306, 751260
Swissprot
expression in Nicotiana tabacum as a protection against tobacco mosaic virus
Swissprot
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
-
-
-
134521, 134522, 134523, 134534, 134537, 134538, 134542, 134543, 134551, 134552, 134553, 134555, 134556, 134558, 134559, 134561, 134562, 134563, 134564, 654993, 655538, 655556, 655907, 656315, 656631, 663931, 665226, 677404, 677546, 678502, 678727, 678732, 678830, 680195, 680196, 681511, 681593, 681707, 682271, 682711, 682714, 682969, 690789, 707863, 717242, 717298, 717371, 717861, 717990, 729310, 729452
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
additional information
-
analysis of synthesis and maturation, folding, quality control, and secretion, of pancreatic RNase in the endoplasmic reticulum of live cells, overview. In contrast to the slow in vitro refolding, the protein folds almost instantly after translation and translocation into the endoplasmatic reticulum lumen. Despite high stability of the native protein, only about half of the RNase reaches a secretion competent, monomeric form and is rapidly transported from the rough endoplasmic reticulum via the Golgi complex to the extracellular space, the rest remains in the endoplasmic reticulum mainly in the form of dimers and is slowly degraded
-
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
RNase A tandem enzymes, in which two RNase A molecules are artificially connected by a peptide linker, and thus have a pseudodimeric structure, exhibit remarkable cytotoxic activity, but can be inhibited by the cytosolic ribonuclease inhibitor in vitro. Structure modeling, overview
physiological function
His12 acts mainly as a general base in the catalytic process of RNase A
evolution
malfunction
-
mechanistic model for the denaturation of bovine pancreatic ribonuclease A in urea, a direct interaction between urea and protonated histidine as the initial step for protein inactivation followed by hydrogen bond formation with polar residues, and the breaking of hydrophobic collapse as the final steps for protein denaturation
physiological function
additional information
evolution
-
the enzyme belongs to the pancreatic-type secretory ribonuclease superfamily
evolution
-
the enzyme belongs to the pancreatic-type secretory ribonuclease superfamily as a unique natively dimeric member
evolution
the enzyme belongs to the vertebrate pancreatic-like RNase A superfamily, sequence comparisons and phylogenetic analysis, overview
physiological function
-
the enzyme has almost no anti-tumoral property in pancreatic adenocarcinoma cell lines and in nontumorigenic cells as normal control, and is largely ineffective as anti-proliferative and pro-apoptotic agent
physiological function
-
the enzyme has good anti-tumoral property in pancreatic adenocarcinoma cell lines and in nontumorigenic cells as normal control, it stimulates a strong anti-proliferative and pro-apoptotic effect in cancer cells. The enzyme triggers Beclin1-mediated autophagic cancer cell death, providing evidence that high proliferation rate of cancer cells may render them more susceptible to autophagy by treatment with the enzyme
additional information
very subtle structural, chemical, and potentially motional variations contribute to ligand discrimination in the enzyme
additional information
-
very subtle structural, chemical, and potentially motional variations contribute to ligand discrimination in the enzyme
additional information
the enzyme performs 3D domain swapping, a process by which two or more protein molecules exchange part of their structure to form intertwined dimers or higher oligomers
additional information
-
the enzyme performs 3D domain swapping, a process by which two or more protein molecules exchange part of their structure to form intertwined dimers or higher oligomers
additional information
-
analysis of synthesis and maturation, folding, quality control, and secretion, of pancreatic RNase in the endoplasmic reticulum of live cells, overview. In contrast to the slow in vitro refolding, the protein folds almost instantly after translation and translocation into the endoplasmatic reticulum lumen. Despite high stability of the native protein, only about half of the RNase reaches a secretion competent, monomeric form and is rapidly transported from the rough endoplasmic reticulum via the Golgi complex to the extracellular space
additional information
-
analysis of the disulfide bond formation phase in detail in the oxidative folding, as the first of two folding phases, of RNase A, overview. Comparision of folding intermediates of reduced RNase A obtained at 25°C and different pH values from pH 4.0, pH 7.0, to pH 10.0, shuffling and transformation of different intermediate types, overview. The preconformational folding phase coupled with disulfide bond formation can be divided into two distinct subphases, a kinetic (or stochastic) disulfide bond formation phase and a thermodynamic disulfide bond reshuffling phase. The transition from kinetically formed to thermodynamically stabilized disulfide bond intermediates are induced by hydrophobic nucleation as well as generation of the native interactions
additional information
-
domain swapping, the process in which a structural unit is exchanged between monomers to create a dimer containing two versions of the monomeric fold, is believed to be an important mechanism for oligomerization and the formation of amyloid fibrils. In RNase residue P114 acts as a conformational gatekeeper, regulating interconversion between monomer and domain-swapped dimer forms, with cis and trans conformation, isomerization at P114 may facilitate population of a partially unfolded intermediate or alternative structure competent for domain swapping, overview
additional information
-
pancreatic ribonuclease A shows domain swapping, a type of oligomerization in which monomeric proteins exchange a structural element, resulting in oligomers whose subunits recapitulate the native, monomeric fold, under extreme conditions, such as lyophilization from acetic acid. The major domain swaps dimer form of RNase A exchanges a beta-strand at its C-terminus to form a C-terminal domain-swapped dimer, mechanism, overview. Domain swapping occurs via a local high-energy fluctuation at the C-terminus
additional information
RNA subsites and reaction mechanism catalyzed by RNase A, molecular interactions between the RNA substrate and residues of the catalytic groove. The following residues are known to interact with each subsite: Lys66 (P0), Thr45 and Asp83 (B1), Gln11, His12, Lys41, His119, and Asp121 (P1), Asn71 and Glu111 (B2), and Lys7 and Arg10 (P2). Structure-function relationship, overview. Potential role for catalytic base His119 in ligand discrimination and/or stabilization in addition to its critical role in catalysis, molecular dynamic simulations show that His119 adopts both rotameric positions in solution, most likely experiencing conformational exchange over the course of a catalytic reaction. Functional importance of long-range conformational rearrangements in RNase A
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). RNAS1_BOVIN
150
0
16461
Swiss-Prot
Secretory Pathway (Reliability: 1)
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
13590
13610
13620
13640
13650
13680
15000
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
dimer
dimer
monomer
oligomer
additional information
dimer
two distinct dimeric forms. In one dimer (AA-CS), the two monmomers swap the C-terminal beta-strand (residues 116-124), while in the other (AA-NS) the two monomers mutually exchange the N-terminal alpha-helix
dimer
dimerization of the P114A mutant enzyme is dimerization is carried out through lyophilization from 40% acetic acid, the mutant efficiently oligomerizes under these conditions, the dimers are separated by gel filtration and ion exchange chromatography
dimer
-
BS-RNase is covalent dimer with two intersubunit disulfide bridges between Cys31 of one chain and Cys32 of the second and vice versa. The native enzyme is an equilibrium mixture of two isomers, MxM and M=M. In the former the two subunits swap their N-terminal helices. In the reducing environment of cytosol, M=M dissociates into monomers, which are strongly inhibited by ribonuclease protein inhibitor, wheras MxM remains as a non-covalent dimer which evades ribonuclease protein inhibitor
dimer
swapping keeps the dimeric structure stable even in the reducing cytosolic environment
dimer
-
besides dimers, also trimers and higher oligomers can be identified among the products of the covalently linking reaction, and the zero-length dimers appear not to be a unique species, but heterogeneous, overview. Quantification of the zero-length oligomers
dimer
-
constituted by two identical subunits covalently bound through two antiparallel disulfide bridges
oligomer
-
ribonuclease A can form a series of 3D domain swapped oligomers by exchanging the N-terminal alpha-helix or the C-terminal beta-strand or both. These oligomers have additional biological and enzymatic activities that the monomeric protein lacks. Ribonuclease A oligomerization is induced by 40% acetic acid, which has been assumed to mildly unfold the protein by detaching the terminal segments and consequently facilitating intersubunit swapping, once the acetic acid is removed by lyophilization and the protein is redissolved in a benign buffer
oligomer
-
the enzyme can oligomerize through the 3D domain swapping of both N- and C-termini, and its multimers is enzymatically and biologically more active than the native dimer
additional information
by lyophilization from 40% acetic acid solutions, bovine pancreatic ribonuclease A forms three-dimensional domain-swapped dimers, trimers, and tetramers. Each oligomeric species consists of at least two conformers, one less basic, one more basic. Identification of a fifth tetramer, pentamers and hexamers, and detection of trace heptameric, octameric and nonameric species
additional information
-
by lyophilization from 40% acetic acid solutions, bovine pancreatic ribonuclease A forms three-dimensional domain-swapped dimers, trimers, and tetramers. Each oligomeric species consists of at least two conformers, one less basic, one more basic. Identification of a fifth tetramer, pentamers and hexamers, and detection of trace heptameric, octameric and nonameric species
additional information
lyophilization of enzyme from 40% acetic acid solution leads to formation of several three-dimensional domain-swapped oligomers, dimers, trimers, tetramers, pentamers. Hexamers, and traces of high-order oligomers. Modeling of tetrameric structure and of larger multimers
additional information
-
lyophilization of enzyme from 40% acetic acid solution leads to formation of several three-dimensional domain-swapped oligomers, dimers, trimers, tetramers, pentamers. Hexamers, and traces of high-order oligomers. Modeling of tetrameric structure and of larger multimers
additional information
bovine pancreatic ribonuclease is able to swap the N-terminal alpha-helix (residues 1-13) and/or the C-terminal beta-strand (residues 116-124), forming a variety of oligomers, including two different dimers. Cis-trans isomerization of the Asn113-Pro114 peptide group is observed when the protein formed the C-terminal swapped dimer. Importance of the hydration shell in determining the cross-talk between the chain termini in the swapping process of RNase A
additional information
-
bovine pancreatic ribonuclease is able to swap the N-terminal alpha-helix (residues 1-13) and/or the C-terminal beta-strand (residues 116-124), forming a variety of oligomers, including two different dimers. Cis-trans isomerization of the Asn113-Pro114 peptide group is observed when the protein formed the C-terminal swapped dimer. Importance of the hydration shell in determining the cross-talk between the chain termini in the swapping process of RNase A
additional information
the reaction between cis-diamminedichloroplatinum(II), cisplatin, a common anticancer drug, and bovine pancreatic ribonuclease, induces extensive protein aggregation, leading to the formation of one dimer, one trimer and higher oligomers whose yields depend on cisplatin/protein ratio. Structural and functional properties of the purified platinated species, together with their spontaneous dissociation and thermally induced denaturation, have been characterized. Platinated species preserve a significant, although reduced, ribonuclease activity. The high resistance of the dimers against dissociation and the different thermal unfolding profiles suggest a quaternary structure different from those of the well-known swapped dimers of RNase A
additional information
-
the reaction between cis-diamminedichloroplatinum(II), cisplatin, a common anticancer drug, and bovine pancreatic ribonuclease, induces extensive protein aggregation, leading to the formation of one dimer, one trimer and higher oligomers whose yields depend on cisplatin/protein ratio. Structural and functional properties of the purified platinated species, together with their spontaneous dissociation and thermally induced denaturation, have been characterized. Platinated species preserve a significant, although reduced, ribonuclease activity. The high resistance of the dimers against dissociation and the different thermal unfolding profiles suggest a quaternary structure different from those of the well-known swapped dimers of RNase A
additional information
-
aggregation of RNaseA might be initiated by hydrophobic interactions, controlled by oligomerization and mediated by electrostatic interactions
additional information
-
enzyme aggregates to form various types of catalytically active oligomers during lyophilization from aqueous acetic acid solution. Each oligomeric species is present in at least two conformational isomers. The review briefly describes the structures of the main oligomers of RNase A, and their principal functional and biological properties
additional information
-
comparison of folding kinetics with Rana pipiens' onconase and bovine angiogenin at pH 8.0 and 15°C. Direct correlation between the number of cis-prolyl bonds in a native protein and the complexity with which it folds via slower phases
additional information
-
construction of enzyme dimer composed of monomeric units covalently linked by a single amide bond between the side-chains of residues K66 and E9 by incubation of a lyophilized preparation of enzyme under vacuum at 85°C. Dimer exhibits a twofold increase in activity over monomeric enzyme and is not inhibited by the cellular ribonuclease inhibitor protein
additional information
-
lyophilization of enzyme from 50% acetic acid solution leads to formation of two dimers and several oligomeric forms. Study on relationship between surface histidine topography in oligomeric forms and catalytic property. Comparison with seminal ribonucleases. Oligomerization also results in modification of the affinity toward the immobilized transition-metal chelate iminodiacetic acid-Cu(II)
additional information
-
protein disulfide isomerase acts both as chaperone and an oxidase during the folding of enzyme. Protein disulfide isomerase catalyzes the conversion of the kinetically trapped enzyme intermediates, des-[26-84] and des-[58-110], by re-shuffling them into the on-pathway intermediate, des-[40-95], and the formation of the native protein
additional information
-
the chemical conjugation of polyethylene glycol to the RNase A C-dimer, and to two trimers, decreases the aspermatogenic activity of the oligomers while increasing their inhibitory activity on the growth of human UB900518 amelanotic melanoma transplanted in athymic nude mice. The conjugated RNase A oligomers are devoid of any embryotoxic activity
additional information
-
oligomers have similar thermal stability to that of monomeric enzyme, suggesting that the main limiting factor in RNase A stability is the tertiary, rather than quaternary structure
additional information
-
ribonuclease A is capable of forming amyloid-like fibrils, the protein is used as a model system to validate fibrillogenic predictions by a 3D profile method based on the crystal structure of the segment NNQQNYand it is demonstrated that a specific residue order is required for fiber formation
additional information
-
the secreted RNase in the bovine pancreas is mainly monomeric, whereas the enzyme present in the cells also contains 20% dimers
additional information
-
native RNase A can be induced to form various N- or C-terminal domain-swapped dimers, trimers, and larger oligomers, all reconstituting the active site and augmenting the enzymatic activity against dsRNA with respect to the native monomer. RNase A oligomers can also acquire cytotoxic activity both in vitro and in vivo
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
glycoprotein
unfolding molecular dynamics simulations of glycosylated and unglycosylated enzyme. Attachment of monomeric N-acetylglucosyamine to residue N34 results in a change of denaturing process. The glycosylated enzyme remains more stable due to preserved non-local interactions
glycoprotein
additional information
-
native enzyme contains four intra-molecular disulfide bonds. Protein disulfide isomerase catalyzes the conversion of the kinetically trapped enzyme intermediates, des-[26-84] and des-[58-110], by re-shuffling them into the on-pathway intermediate, des-[40-95], and the formation of the native protein
glycoprotein
-
serum ribonucleases differ in glycosylation, extent varies from day to day in an individual
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, crystal structure of the cis-Pro to Gly variant P114G, structure solved at 2.0 A resolution, space group: P4(3)2(1)2
bovine pancreatic ribonuclease A is crystallized from a mixture of small molecules containing basic fuchsin, tobramycin and uridine 5-monophosphate. Solution of the crystal structure reveals that the enzyme is selectively bound to uridine 5-monophosphate, with the pyrimidine ring of uridine 5-monophosphate residing in the pyrimidine-binding site at Thr45, description of the mode of binding of the nucleotide to the enzyme, crystal structure of bovine pancreatic ribonuclease complexed with uridine-5-monophosphate at 1.60 A resolution, 0.1 M HEPES buffer, reservoir of 25% PEG3350 in water, droplets are 5-10 mM in basic fuchsin, tobramycin, uridine-5-monophosphate and 1 mM in ribonuclease A, pH 7.0, vapor diffusion, sitting drop, temperature 298 K
explicit-solvent molecular dynamics simulations up to the melting temperature of 64°C. Between 37°C and 47°C, there is a small but significant decrease in the number of native contacts, beta-sheet hydrogen bonding, and deviation of backbone conformation, and an increase in the number of non-native contacts. At 57°C°C and 67°C, a non-native helical segment of residues 15-20 forms
hanging drop vapor diffusion method, ammonium sulfate, sodium chloride, sodium acetate, pH 6.0, crystal structure of bovine pancreatic ribonuclease A (wild-type), resolution 1.60 A, crystal structure of bovine pancreatic ribonuclease A variant V47A, resolution 1.60 A, crystal structure of bovine pancreatic ribonuclease A variant V54A, resolution 1.60 A, crystal structure of bovine pancreatic ribonuclease A variant V57A, resolution 1.60 A, crystal structure of bovine pancreatic ribonuclease A variant I81A, resolution 2.0 A, crystal structure of bovine pancreatic ribonuclease A variant I106A, resolution 1.40 A, crystal structure of bovine pancreatic ribonuclease A variant V108A, resolution 1.60 A, space group P3221
hanging-drop/vapor-diffusion method, 20% PEG 4000, 0,02 M sodium citrat buffer, pH 5.5, 16°C, pancreatic ribonuclease A-5-ATP complex, resolution 1.70 A, pancreatic ribonuclease A-P3-bis(5-adenosyl) triphosphate complex, resolution 2.40 A, pancreatic ribonuclease A-NADPH complex, resolution 1.70 A, pancreatic ribonuclease A-NADP complex, resolution 1.70 A, pancreatic ribonuclease A-pyrophosphte ion complex, resolution 1.80 A, space group C121
in complex with inhibitor 3-N-piperidine-4-carboxyl-3-deoxy-ara-uridine at 1.7 A resolution. Two inhibitor molecules bind in the central cavity of enzyme, the first occupying the purine-preferring site, and the second molecule binding to the carboxyl group at the pyrimidine recognition site
multi-ns molecular dynamics simulations of enzyme in complex with inhibitor 5'-phospho-2'-deoxyuridine-3-pyrophosphate (P-5)-adenosine-3-phosphate. The adenylate 5'-beta-phosphate binding position and the adenosine syn orientation constitute robust structural features in the complex
multiple solvent crystal structures, vapor diffusion, hanging drop, PEG 4000, sodium citrate, pH 5.0, temperature 291 K, crystal structure of crosslinked ribonuclease A, resolution 1.65 A, crosslinked crystals are then transferred with a cryo-loop to new drops containing stabilization buffer and an organic solvent and allowed to soak for 1-2 h at room temperature. Soaked crystals are then collected, cryo-protected by dunking in stabilization buffer containing 20% glycerol, and flash frozen in liquid nitrogen, crystal structure of ribonuclease A in 50% dimethylformamide, resolution 1.84 A, crystal structure of ribonuclease A in 50% dioxane, resolution 1.95 A, crystal structure of ribonuclease A in 70% dimethyl dulfoxide, resolution 1.76 A, crystal structure of ribonuclease A in 70% 1,6-hexanediol, resolution 2.00 A, crystal structure of ribonuclease A in 70% isopropanol, resolution 2.02 A, crystal structure of ribonuclease A in 70% t-butanol, resolution 1.68 A, crystal structure of ribonuclease A in 50% trifluoroethanol, resolution 1.93 A, crystal structure of ribonuclease A in 1 M trimethylamine N-oxide, resolution 1.68 A, crystal structure of ribonuclease A in 50% R,S,R-bisfuranol, resolution 1.76 A, comparison of the multiple solvent crystal structures with inhibitor-bound crystal structures of ribonuclease A reveals that the organic solvent molecules identify key interactions made by inhibitor molecules, highlighting ligand binding hot-spots in the active site, investigation of plasticity, hydration and clustering of organic solvent molecules in the active site
mutant V43C/R85C at 1.6 A resolution. Residues V43 and R85 are not involved in the folding/unfolding transition states ensemble, and residues A4 and V118 may form non-native contacts
neutron crystallographic analysis of phosphate-free bovine pancreatic RNase A, 50% tert-butyl alcohol, temperature 298 K, then the crystal is soaked in heavy water solution, pH 6.2, for two months, BATCH, space group P1211, resolution 1.7 A, His12 acts mainly as a general base in the catalytic process of Rnase A, numerous other distinctive structural features such as the hydrogen positions of methyl groups, hydroxyl groups, prolines, asparagines and glutamines are also determined
purified enzyme mutant P114A in monomeric and dimeric form, for the monomeric enzyme hanging drop vapour diffusion method is used mixing of 24 mg/ml protein with reservoir solution containing 35% w/v ammonium sulfate, 50% v/v of saturated NaCl, and 0.1 M acetate buffer, pH 6.6, 20°C, 1 week, for the dimeric enzyme sitting drop vapour diffusion method is used with 15 mg/ml protein mixed with precipitation solution containing 17-19% w/v PEG 20000, 0.1 M cacodylate buffer, pH 6.5, 100-150 mg/ml of trehalose, and 11 mM of 2'-deoxycytidylyl(3',5')-2'-deoxyguanosine, a few days, 20°C, X-ray diffraction structure determination and analysis at 2.10 A and 2.18 A resolution, respectively, molecular replacement
purified platinated monomeric enzyme, obtained upon RNase A incubation in 1:10 protein to metallodrug ratio, is crystallized at 25°C using the hanging drop vapor diffusion method
ribonuclease A in complex with thymidine 3'-monophosphate, hanging drop vapor diffusion method, 0.002 ml of 80 mg/ml protein in 20% ethanol and 20% acetic acid at pH 5.5, is mixed with 0.004 ml of 3'-TMP dissolved in a mother liquor solution of 20% ammonium sulfate and 2 M sodium chloride at pH 5.5, room temperature, 1 week, X-ray diffraction structure determination and analysis at 1.55 A resolution, molecular replacement, modelling
RNase A tandem enzymes, hanging drop vapor diffusion method, mixing of 0.002 ml of 10 mg/ml protein in 10 mm Tris-HCl, pH 7.0, with 0.002 ml of reservoir solution containing 30% w/v PEG 8000 and 200 mm (NH4)2SO4, 6 days, 13°C, X-ray diffraction structure determination and analysis at 1.68 A resolution
structural investigation of ribonuclease A conformational preferences using high pressure protein crystallography
vapor diffusion, hanging drop, PEG 4000, sodium citrate, pH 5.5, 289 K, space group C121,ribonuclease A-1-{5-deoxy-5-[4-(ethoxycarbonyl)piperidin-1-yl]-alpha-L-arabinofuranosyl}pyrimidine-2,4(1H,3H)-dione complex, resolution 1.58 A, ribonuclease A-1-(5-deoxy-5-morpholin-4-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione, resolution 1.60 A, ribonuclease A-1-(5-deoxy-5-piperidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione, resolution 1.60 A, ribonuclease A-1-(5-deoxy-5-pyrrolidin-1-yl-alpha-L-arabinofuranosyl)pyrimidine-2,4(1H,3H)-dione, resolution 1.60 A, ribonuclease A-5-deoxy-5-piperidin-1-ylthymidine, resolution 1.72 A, ribonuclease A-1-(2,5-dideoxy-5-pyrrolidin-1-yl-beta-L-erythro-pentofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione, resolution 1.98 A
vapor-diffusion, hanging-drop, PEG 4000, sodium citrate, pH 5.5, temperature 289 K, ribonuclease A-uridine 5 phosphate complex, resolution 1.39 A, space group C121, ribonuclease A-uridine 5 diphosphate complex, resolution 1.40 A, space group C121
at 1.33 A resolution, space group P31. Structure contains two molecules of nucleotide per enzyme molecule, one in the active site cleft in the productive binding mode, the other occupies the pyrimidine-specific binding site in a non-productive mode
-
circular dichroism study on the conformation of enzyme in a miniemulsion. The addition of poly(vinyl alcohol) as a co-surfactant is effective in preserving the protein structural integrity
-
comparison of mutant crystal structures, PDB IDs 3RSK, 1A5P, and 1C9V, with the wild-type structure, PBD ID 1FS3, overview
-
crystallization in presence of 2'-deoxycitidylyl(3'-5')-2'-deoxyadenosine at 4°C by using sitting drop vapor diffusion method. Crystal structure of the MxM isomer of the enzyme in the non-covalent dimer form, carboxyamidomethylated at residues Cys31 and Cys32, in a complex with 2'-deoxycitidylyl(3'-5')-2'-deoxyadenosine
-
data of enzyme dimer composed of monomeric units covalently linked by a single amide bond between the side-chains of residues K66 and E9 by incubation of a lyophilized preparation of enzyme under vacuum at 85°C. Procedure does not induce a significant conformational change
-
pressure tuning hole burning experiments using the UV-absorbing tyrosine residues. Ribonuclease A protein stays intact upon cooling to 2 K. Its various tyrosine sites show characteristic features which can be resolved in pressure tuning hole burning spectra. Reducing the sulfur bridges leads to a loss of the individual features, and the sites become alike. The respective compressibility is reduced by more than a factor of 2 and comes close to the value of free tyrosine in solution. Compared to the reduction of the sulfur bridges, the influence of guanidinium hydrochloride on the pressure tuning behavior is less pronounced
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
A20P
thermodynamic and kinetic stability is similar to wild-type ribonuclease A. Mutation has no significant effect on the native conformation and catalytic activity
A20P/S21P
thermodynamic and kinetic stability is similar to wild-type ribonuclease A. Mutation has no significant effect on the native conformation and catalytic activity
D121A
with nearly the same Km value as the wild type enzyme but with lower hydrolytic activity
D121A/S123A/V124A
mutant in which all of the C-terminal four amino acid residues are replaced by alanine residues, with a final lower hydrolytic activity
D121E
with nearly the same Km value as the wild type enzyme but with with lower hydrolytic activity
D121K
with nearly the same Km value as the wild type enzyme but with with lower hydrolytic activity
DELTA121-124
C-terminal deletion mutant, 14ºC less stable to thermal denaturation than the wild type enzyme
DELTA122-124
C-terminal deletion mutant, 3-6ºC less stable to thermal denaturation than the wild type enzyme
DELTA123-124
C-terminal deletion mutant, 3-6ºC less stable to thermal denaturation than the wild type enzyme
DELTA124
C-terminal deletion mutant, 3-6ºC less stable to thermal denaturation than the wild type enzyme
F120A
F46A
F46L
F46V
mutant with adversely affected conformational stability and folding speed
F46Y
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
F8A
substitution of Phe8 results in a recombinant variant significantly destabilized
F8L
substitution of Phe8 results in a recombinant variant significantly destabilized
I106A
I81A
K31A/R33S
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
K31A/R33S/F46Y
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
K7H/R10H
17% of wild-type activity, introduction of a putative new catalytic site resulting in increase in exonucleolytic activity
K7H/R10H/H12K/H119Q
9% of wild-type activity due to suppression of native active site, increase in exonucleolytic activity
L35A
L35A/F46Y
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
L35S
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
L35S/F46Y
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
L51A
shortening the side chain of the hydrophobic solvent-exposed residue Leu51 to Ala has almost no effect in the stability of ribonuclease A
N121X
L-alpha-Asp at position 121 in RNase A is replaced by L-beta-, D-alpha-, and D-beta-Asp. The objective aspartic acid at position 121 is located near the active site and related to RNA cleavage. The RNase A with L-alpha-Asp at position 121 shows a normal activity. The catalytic activity of L-beta-, D-alpha-, and D-beta-Asp-containing RNase A is markedly decreased
N34D
thermodynamic and kinetic stability is similar to wild-type ribonuclease A. Mutation has no significant effect on the native conformation and catalytic activity
P114A
site-directed mutagenesis, the mutation at the C-terminus affects the capability of the N-terminal alpha-helix to swap and the stability of both dimeric forms
P114G
three hydrogen bonds and two bifurcated hydrogen bonds present in the cis wild-type structure are replaced by four hydrogen bonds and two bifurcated hydrogen bonds in the P114G structure
Q28A
promotes an increase in the helix propensity, from 0.99 in the wild-type to 1.5
S21L
thermodynamic and kinetic stability is similar to wild-type ribonuclease A. Mutation has no significant effect on the native conformation and catalytic activity
S21P
thermodynamic and kinetic stability is similar to wild-type ribonuclease A. Mutation has no significant effect on the native conformation and catalytic activity
V108A
V43A
substitution of the hydrophobic residue valin 43 by alanin results in an increase in the global stability of the ribonuclease A structure of 4.02 kJ/mol in free energy
V43C/R85C
V47A
V54A
V57A
C40A/C95A
G38K
-
the mutant is more basic and interacts more strongly with the acidic membrane of cancer cells compared to the wild-type enzyme
H12A
-
comparison of mutant crystal structure, PDB ID 1C9V, with the wild-type structure, PBD ID 1FS3
I106A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
I107A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
I81A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
K31C/S32C/S16G/T17N/A19P/A20S
site-directed mutagenesis, very poor cytotoxic activity
K31C/S32C/S16G/T17N/A19P/A20S/S80R
site-directed mutagenesis, very poor cytotoxic activity
K66A
-
site-directed mutagenesis, no intramolecular bonds form in the K66A variant
K7A/R10A/K66A
-
comparison of mutant crystal structure, PDB ID 3RSK, with the wild-type structure, PBD ID 1FS3
L35M
-
unchanged in respect to folding and stability, but with enhanced glycosylation
N113S
-
the mutant N113S is more prominent in the Golgi than wild-type bovine RNase, which is mainly present in the endoplasmic reticulum
P114A
-
site-directed mutagenesis, the mutant adopts a trans conformation in contrast to the wild-type which shows a cis conformation
P114G
P93A
T17A
-
site-directed mutagenesis, the mutant shows reduced affinity and binding to inhibitor 3'-CMP compared to the wild-type enzyme, kinetics, and conformational exchange motions, overview
T82A
-
site-directed mutagenesis, the mutant shows reduced affinity and binding to inhibitor 3'-CMP compared to the wild-type enzyme, kinetics, and conformational exchange motions, overview
V108A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
V124A
-
C-terminus involved in the formation of disulfide bonds during refolding process
V124E
-
C-terminus involved in the formation of disulfide bonds during refolding process
V124G
-
C-terminus involved in the formation of disulfide bonds during refolding process
V124K
-
C-terminus involved in the formation of disulfide bonds during refolding process
V124L
-
C-terminus involved in the formation of disulfide bonds during refolding process
V124W
-
C-terminus involved in the formation of disulfide bonds during refolding process
V47A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
V54A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
V57A
-
thermodynamic analysis of pressure-unfolding and kinetics for positive pressure-jumps
Y115W
additional information
F46A
mutant with adversely affected conformational stability and folding speed
F46L
mutant with adversely affected conformational stability and folding speed
L35A
thermodynamic and kinetic stability of the mutant is greatly decreased. Mutation has no significant effect on the native conformation and catalytic activity
V43C/R85C
crystallization data. Residues V43 and R85 are not involved in the folding/unfolding transition states ensemble
C40A/C95A
-
comparison of mutant crystal structure, PDB ID 1A5P, with the wild-type structure, PBD ID 1FS3
P114G
-
site-directed mutagenesis, the mutant adopts a trans conformation in contrast to the wild-type which shows a cis conformation
P114G
-
site-directed mutagenesis, the mutant adopts a trans conformation in contrast to the wild-type who shows a cis conformation. The P114G mutant readily domain swaps under physiological conditions in contrast to the wild-type enzyme. The P114G variant has decreased protection from hydrogen exchange compared to the wild-type protein near the C-terminal hinge region. Structure of RNase A P114G with HX fluctuation, overview
Y115W
-
large increase in fluorescence yield upon unfolding. Analysis of reversible pressure dependent unfolding profiles. With increasing temperature, the sigmoidal unfolding transition is shifted towards higher pressures
additional information
A4C/G88R/V118C mutant, with a new disulfide bond that links the N and C termini, is more stable to thermal denaturation than wild type and G88R enzymes. The conformational stability of the C40A/G88R/C95A and C65A/C72A/G88R mutants is less than that of G88R. A4C/C65A/C72A/G88R/V118C mutant, with a new disulfide bond is more stable than C65A/C72A/G88R mutant
additional information
-
A4C/G88R/V118C mutant, with a new disulfide bond that links the N and C termini, is more stable to thermal denaturation than wild type and G88R enzymes. The conformational stability of the C40A/G88R/C95A and C65A/C72A/G88R mutants is less than that of G88R. A4C/C65A/C72A/G88R/V118C mutant, with a new disulfide bond is more stable than C65A/C72A/G88R mutant
additional information
series of mutations in residues V54, V57, I106 and V108 most critical for stability. Detailed study on thermodynamic parameters
additional information
transgenic expression in Nicotiana tabacum as a protection against tobacco mosaic virus. Transgenic plants are characterized by an increased level of enzyme activity in leaf extract and exhibit a significantly higher level of protection against the virus infection than control. Protection is evident by the absence or significant delay of the appearance of typical mosaic symptoms and the retarded accumulation of infectious virus and viral antigen
additional information
-
transgenic expression in Nicotiana tabacum as a protection against tobacco mosaic virus. Transgenic plants are characterized by an increased level of enzyme activity in leaf extract and exhibit a significantly higher level of protection against the virus infection than control. Protection is evident by the absence or significant delay of the appearance of typical mosaic symptoms and the retarded accumulation of infectious virus and viral antigen
additional information
use of gene duplication to generate tandem enzymes covalently bound by peptide linker. Tandemization has minor effects on the activity and stability in comparison to monomeric RNase A. Relative activity decreases by 10-50%, and melting temperature decreases by less than 2.5 K. Tandemization results in remarkable cytotoxicity, decreasing the IC50 values with K-562 cells to 0.070-0.013 mM
additional information
-
use of gene duplication to generate tandem enzymes covalently bound by peptide linker. Tandemization has minor effects on the activity and stability in comparison to monomeric RNase A. Relative activity decreases by 10-50%, and melting temperature decreases by less than 2.5 K. Tandemization results in remarkable cytotoxicity, decreasing the IC50 values with K-562 cells to 0.070-0.013 mM
additional information
the structure of bovine pancreatic ribonuclease A variants V47A, V54A, V57A, I81A, I106A, and V108A is solved at 1.4-2.0 A resolution and compared with the structure of wild-type protein. The introduced mutations have only minor influence on the global structure of ribonuclease A. The structural changes have individual character that depends on the localization of mutated residue, however, they seem to expand from mutation site to the rest of the structure. Analysis of the difference distance matrices reveal that the ribonuclease A molecule is organized into five relatively rigid subdomains with individual response to mutation
additional information
-
the structure of bovine pancreatic ribonuclease A variants V47A, V54A, V57A, I81A, I106A, and V108A is solved at 1.4-2.0 A resolution and compared with the structure of wild-type protein. The introduced mutations have only minor influence on the global structure of ribonuclease A. The structural changes have individual character that depends on the localization of mutated residue, however, they seem to expand from mutation site to the rest of the structure. Analysis of the difference distance matrices reveal that the ribonuclease A molecule is organized into five relatively rigid subdomains with individual response to mutation
additional information
-
construction of enzyme dimer composed of monomeric units covalently linked by a single amide bond between the side-chains of residues K66 and E9 by incubation of a lyophilized preparation of enzyme under vacuum at 85°C. Procedure does not induce a significant conformational change, dimer shows an 2fold increase in activity over monomeric enzyme and is not inhibited by the cellular ribonuclease inhibitor protein
additional information
-
cross-linking of enzyme or its covalently linked dimer to polyspermine using dimethyl suberimidate. The in vitro and in vivo cytotoxic activity of treated monomeric enzyme is not higher than that of free polyspermine, but dimeric suberimidate-treated enzyme proves to be a more efficient antitumor agent both in vitro and in vivo
additional information
-
substitution of P114 with residues that strongly prefer a trans peptide bond, like Ala, Gly, results in significant population of the C-terminal domain-swapped dimer under near-physiological conditions of pH 8.0 and 37°C. This is in stark contrast to dimerization of wild-type RNase A, which requires incubation under extreme conditions such as lyophilization from acetic acid or elevated temperature
additional information
-
two RNase A variants, P114G and P93A, have the same global stability yet very different domain-swapping propensity, differences in protection factors suggest differential local dynamics
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 9
-
maximum conformational stability to urea and guanidine hydrochloride denaturation
24208
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45 - 66
thermal denaturation temperatures Tm of enzyme mutant P114A, wild-type enzyme RNase A, and N- and C-swapped dimers of the two proteins, overview
47.7
35 - 40
-
outer shell of ribonuclease structure begins to unfold, steps in pathway of thermal unfolding
40 - 70
-
oligomers have similar thermal stability to that of monomeric enzyme, suggesting that the main limiting factor in RNase A stability is the tertiary, rather than quaternary structure
65
-
antibodies against native RNase or against RNase N-terminal dodecapeptide are effective in lowering aggregation at 65°C
85
-
thermal denaturation of RNase A, alone or in the presence of cationic gemini surfactants, is reversible
additional information
additional information
des-124, des-(123-124), des-(122-124) mutants are 3-6°C less stable to thermal denaturation with respect to the wild type enzyme. des-(121-124) mutant is 14°C less stable to thermal denaturation with respect to the wild type enzyme
additional information
-
des-124, des-(123-124), des-(122-124) mutants are 3-6°C less stable to thermal denaturation with respect to the wild type enzyme. des-(121-124) mutant is 14°C less stable to thermal denaturation with respect to the wild type enzyme
additional information
two distinct dimeric forms. In one dimer (AA-CS), the two monmomers swap the C-terminal beta-strand (residues 116-124), while in the other (AA-NS) the two monomers mutually exchange the N-terminal alpha-helix. Thermal denaturation of both dimers can be described by a two-step dissociation/unfolding mechanism. The structural determinants for the higher stability of AA-NS should reside in its open interface
additional information
explicit-solvent molecular dynamics simulations up to the melting temperature of 64°C. Between 37°C and 47°C, there is a small but significant decrease in the number of native contacts, beta-sheet hydrogen bonding, and deviation of backbone conformation, and an increase in the number of non-native contacts. At 57°C and 67°C, a non-native helical segment of residues 15-20 forms
additional information
-
explicit-solvent molecular dynamics simulations up to the melting temperature of 64°C. Between 37°C and 47°C, there is a small but significant decrease in the number of native contacts, beta-sheet hydrogen bonding, and deviation of backbone conformation, and an increase in the number of non-native contacts. At 57°C and 67°C, a non-native helical segment of residues 15-20 forms
additional information
heat and pressure produce only slightly different energetic changes in the unfolded state of enzyme. Stability differences in mutants can be attributed to both hydrophobic interactions and packing density
additional information
The thermal stability of ribonuclease A and its variants is investigated by circular dichroism spectroscopy measurements. Thermal unfolding proves to be reversible and follows a two-state transition model as judged from the fit of the data. While H105C/V124C-ribonuclease A, the variant with the disulfide bond homolog to onconase is as stable as ribonuclease A and A4C/V118C- and V43C/R85C-ribonuclease A are more stable than ribonuclease A, R10C/R33C-, I107C/A122C-, and particularly M30C/N44C-ribonuclease A are destabilized in comparison to ribonuclease A
additional information
-
His12 mutants enzymes show low thermal stability, with a Tm of 45ºC. Wild type enzyme and His119 mutant show a Tm of 61ºC
additional information
-
type I isoenzyme, Tm of 59°C. Type II isoenzyme, Tm of 53°C
additional information
-
4-chlorobutan-1-ol induces reversible thermal transition in ribonuclease A at low concentrations, irreversible at intermediate concentrations (50-250 mM) and again reversible transitions at further higher concentrations of the alcohol (250-400 mM)
additional information
-
analysis of conformational changes by picosecond time-resolved fluorescence of the six tyrosine residues. Upon thermal or chemical unfolding only Y25, Y92, and Y76 undergo significant displacement from their nearest -SS- bridge. A single unfolding event around 59°C affects all these residues similarly
additional information
-
analysis of thermal unfolding shows a step-wise thermal denaturation mechanism in which the structural adjustment of the N-terminal and the opening of the central structure come before the main unfolding process. Non-native turns form along with the unfolding of the native structures. The central region is the most stable part
additional information
-
Fourier transformation study of thermal denaturation in D2O solution, using sample-sample two-dimensional correlation spectroscopy and principal component analysis. Enzyme undergoes a pretransition at 46°C as well as a main transition at 66°C
additional information
-
L-arginine suppresses heat-induced deamidation and beta-elimination at 98°C, resulting in the suppression of thermal inactivation of bovine pancreas ribonuclease A. Poly(ethylene glycol) with molecular weight 1000 acts as a thermoinactivation suppressor for the protein, especially at higher protein concentrations, while Arg was not effective at higher protein concentrations. Amphiphilic poly(ethylene glycol) and poly(vinylpyrolidone) inhibit intermolecular collision of thermally denatured proteins by preferential interaction with thermally denatured proteins, resulting in the inhibition of intermolecular disulfide exchange, molecular mechanism, overview
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
calculation of thermodynamic parameters from thermally induced unfolding curves
two distinct dimeric forms. In one dimer (AA-CS), the two monmomers swap the C-terminal beta-strand (residues 116-124), while in the other (AA-NS) the two monomers mutually exchange the N-terminal alpha-helix. The two dimers are metastable and dissociate spontaneously to monomers with different kinetics
4-chlorobutan-1-ol induces reversible thermal transition in ribonuclease A at low concentrations, irreversible at intermediate concentrations (50-250 mM) and again reversible transitions at further higher concentrations of the alcohol (250-400 mM)
-
oligomers have similar thermal stability to that of monomeric enzyme, suggesting that the main limiting factor in RNase A stability is the tertiary, rather than quaternary structure
-
urea, denaturation, urea interferes with interhydrophobic interactions by affecting the water molecules
-
urea, low concentrations of salts, CaCl2, LiClO4 or LiCl, stabilize against urea denaturation at higher concentration they destabilize
-
ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
acetic acid
lyophilization of enzyme from 40% acetic acid solution leads to formation of several three-dimensional domain-swapped oligomers, dimers, trimers, tetramers, pentamers. Hexamers, and traces of high-order oligomers. Modeling of terameric structure and of larger multimers
guanidine-HCl
real-time photo-CIDNP spectra from guanidine-HCl refolding experiments
urea
photo-CIDNP NMR spectroscopy for monitoring of the real-time refolding of ribonuclease A following dilution from a high concentration of urea denaturant
urea
-
no helical structure is found in 8 M urea at pH 2.5, ribonuclease A can oligomerize after thorough unfolding in concentrated solutions of urea, followed by a gel filtration step, which exchanges the denaturant for a refolding buffer. The yield of ribonuclease A oligomers depends on the logarithm of ribonuclease A concentration during refolding.
additional information
additional information
-
The conformation of ribonuclease A in 40% acetic acid is found to be mostly but not completely unfolded. All X-Pro bonds are predominantly in the trans conformation and the hydrophobic core and the beta-sheet structure unfold completely. However, three alpha-helices are partly populated in ribonuclease A in 40% acetic acid in approximately the same positions as the three native helices.
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purification of the covalent RNase A oligomers by two-step cation exchange chromatography and two gel filtration steps
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
expression in Escherichia coli strain BL21
expression in Neurospora crassa, seven different vectors are constructed that vary in the use of promoters, open reading frames, and terminators, ccg1 and cfp promoters provide superior efficiency for recombinant gene expression
expression in Nicotiana tabacum as a protection against tobacco mosaic virus
expression in Escherichia coli
expression of the fusion protein with the minor coat protein of phage M13 in M13
-
RENATURED/Commentary
ORGANISM
UNIPROT
LITERATURE
folding/unfolding kinetics of enzyme with and without a covalent crosslink in the form of a fifth disulfide bond. Residues V43 and R85 are not involved in the folding/unfolding transition states ensemble, and residues A4 and V118 may form non-native contacts
photo-CIDNP NMR spectroscopy study for monitoring of the real-time refolding of ribonuclease A following dilution from a high concentration of urea or guanidine-HCl denaturant, two distinct kinetic processes are apparent, a faster step with time constant of 4-8 s (4.2 s in urea and 7.3/7.6 s in guanidine-HCl) and a slower one with time constant 16-24 s (16.6 s in urea and 24.0 s in guanidine-HCl). The latter is attributed to the cis-trans isomerization of Pro 93 and is responsible for the slow disappearance of the signal from Tyr 92 in the folding intermediate observed in the guanidine-HCl experiment
unfolding molecular dynamics simulations of glycosylated and unglycosylated enzyme. Attachment of monomeric N-acetylglucosyamine to residue N34 results in a change of denaturing process. The glycosylated enzyme remains more stable due to preserved non-local interactions
4-chlorobutan-1-ol induces reversible thermal transition in ribonuclease A at low concentrations, irreversible at intermediate concentrations (50-250 mM) and again reversible transitions at further higher concentrations of the alcohol (250-400 mM)
-
analysis of conformational changes by picosecond time-resolved fluorescence of the six tyrosine residues. Upon thermal or chemical unfolding only Y25, Y92, and Y76 undergo significant displacement from their nearest -SS- bridge. A single unfolding event around 59°C affects all these residues similarly
-
analysis of reversible pressure dependent unfolding profiles of mutant Y115W. With increasing temperature, the sigmoidal unfolding transition is shifted towards higher pressures
-
comparison of folding kinetics with Rana pipiens' onconase and bovine angiogenin at pH 8.0 and 15°C. Direct correlation between the number of cis-prolyl bonds in a native protein and the complexity with which it folds via slower phases
-
protein disulfide isomerase acts both as chaperone and an oxidase during the folding of enzyme. Protein disulfide isomerase catalyzes the conversion of the kinetically trapped enzyme intermediates, des-[26-84] and des-[58-110], by re-shuffling them into the on-pathway intermediate, des-[40-95], and the formation of the native protein
-
refolding of denatured and reduced RNase A with refolding-facilitating media immobilized with three folding machineries, mini-chaperone (a monomeric apical domain consisting of residues 191-345 of GroEL) and two foldases (DsbA and human peptidyl-prolyl cis-trans isomerase) by mimicking oxidative refolding chromatography. For efficient and simple purification and immobilization simultaneously, folding machineries are fused with the positively-charged consecutive 10-arginine tag at their C-terminal. The immobilized folding machineries are fully functional when assayed in a batch mode
-
study on enzyme folding and refolding kinetics using pressure-jump techniques. The structure of the transition state is a relatively uniformly expanded form half-way between the folded and unfoded states. The pressure-folding transition state looks like a collapsed globule with some secondary structure and a weakenend hydrophobic core
-
study on folding/unfolding by pressure-jump-induced relaxation kinetics. Downward pressure jumps result always in single exponential kinetics, while upward jumps are biphasic in the low pressure range and monophasic at higher pressures. Analysis of the activation volume shows a temperature-dependent shift of the unfolding transition state to a larger volume
-
the ribonuclease A equilibrium unfolding in urea and guanidinium chloride solutions proceeds through a formation of intermediates whose compactness, retention of the larger part hydrophobic core, secondary structure, and native-like folding pattern correspond to the wet molten globule state. The urea intermediate is less compact than that in GuCl. The refolding of the protein denatured by GuCl results in the formation of the intermediate which enzyme activity is virtually the same as the activity of the native protein
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
agriculture
transgenic expression in Nicotiana tabacum as a protection against tobacco mosaic virus. Transgenic plants are characterized by an increased level of enzyme activity in leaf extract and exhibit a significantly higher level of protection against the virus infection than control. Protection is evident by the absence or significant delay of the appearance of typical mosaic symptoms and the retarded accumulation of infectious virus and viral antigen
medicine
use of gene duplication to generate tandem enzymes covalently bound by peptide linker. Tandemization has minor effects on the activity and stability in comparison to monomeric RNase A. Relative activity decreases by 10-50%, and melting temperature decreases by less than 2.5 K. Tandemization results in remarkable cytotoxicity, decreasing the IC50 values with K-562 cells to 0.070-0.013 mM
pharmacology
inhibitors can be the starting point for the development of compounds that can be used as pharmaceuticals against pathologies associated with ribonuclease A homologues such as human angiogenin, which is implicated in tumor induced neovascularization
analysis
biotechnology
-
synthesis of molecularly imprinted polymers from the monomers styren and polyethyleneglycol 400 dimethacylate with high rebinding efficiency of RNase A to polymer. Polymers show high selectivity for RNase A and high stability
medicine
pharmacology
-
radical-scavenging effects of the ribonuclease inhibitor CPRI may contribute to its function in the cell protection from peroxidative injuries unrelated to inhibition of RNases
analysis
-
preparation of polymeric nanoparticles imprinted with RNase A via miniemulsion polymerization using methyl methacrylate and ethylene glycol dimethacrylate. The addition of poly(vinyl alcohol) as a co-surfactant is effective in preserving the protein structural integrity. Imprinted nanoparticles produces by the optimized method show increased target specificity
analysis
-
pressure tuning hole burning experiments using the UV-absorbing tyrosine residues. Ribonuclease A protein stays intact upon cooling to 2 K. Its various tyrosine sites show characteristic features which can be resolved in pressure tuning hole burning spectra. Reducing the sulfur bridges leads to a loss of the individual features, and the sites become alike. The respective compressibility is reduced by more than a factor of 2 and comes close to the value of free tyrosine in solution. Compared to the reduction of the sulfur bridges, the influence of guanidinium hydrochloride on the pressure tuning behavior is less pronounced
analysis
-
study on spermidine modulation of enzyme activity via individual RNA plasmon rulers which combine high throughput with high temporal resolution at the single molecule level and are able to retrieve otherwise obscured information about weak structural stabilizations
analysis
-
synthesis of molecularly imprinted polymers from the monomers styren and polyethyleneglycol 400 dimethacylate with high rebinding efficiency of RNase A to polymer. Polymers show high selectivity for RNase A and high stability
analysis
-
synthesis of monodispersed RNase A surface-imprinted particles with good magnetic property for practical bioseparation. Use of methyl methacrylate and ethylene glycol dimethacrylate as the functional and cross-linker monomers to produce particles of 700-800 nm diameter imprinted with ribonuclease A and encapsulated with nanosized Fe3O4 particles. Surface-imprinted particles show good selectivity toward the RNase template over control protein
medicine
-
cross-linking of enzyme or its covalently linked dimer to polyspermine using dimethyl suberimidate. The in vitro and in vivo cytotoxic activity of treated monomeric enzyme is not higher than that of free polyspermine, but dimeric suberimidate-treated enzyme proves to be a more efficinet antitumor agent both in vitro and in vivo
medicine
-
the chemical conjugation of polyethylene glycol to the RNase A C-dimer, and to two trimers, decreases the aspermatogenic activity of the oligomers while increasing their inhibitory activity on the growth of human UB900518 amelanotic melanoma transplanted in athymic nude mice. The conjugated RNase A oligomers are devoid of any embryotoxic activity
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Pace, C.N.; Laurents, D.V.; Thomson, J.A.
pH Dependence of the urea and guanidine hydrochloride denaturation of ribonuclease A and ribonuclease T1
Biochemistry
29
2564-2572
1990
Bos taurus
Richards, F.M.; Wyckoff, H.W.
Bovine pancreatc ribonuclease
The Enzymes, 3rd Ed. (Boyer, P. D. , ed. )
4
647-806
1971
Bos taurus
-
Blackburn, P.; Moore, S.
Pancreatic ribonuclease
The Enzymes, 3rd Ed. (Boyer, P. D. , ed. )
15
317-433
1982
Bos taurus
-
Anfinsen, C.B.; White, F.H.
The ribonucleases: occurence, structure and properties
The Enzymes, 2nd Ed. (Boyer, P. D. ; Lardy, H. ; Myrbck, K. , eds. )
5
95-122
1961
Bos taurus, Homo sapiens, Mus musculus, Rattus norvegicus, Sus scrofa
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Fink, A.L.; Anderson, W.D.; Hattersley, J.E.; Lustig, B.S.
The effect of methanol and temperature on the kinetics of refolding of ribonuclease A
FEBS Lett.
236
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1988
Bos taurus
Corbishley, T.P.; Johnson, P.J.; Williams, R.
Serum ribonuclease
Methods Enzym. Anal. , 3rd Ed. (Bergmeyer, H. U. , ed. )
4
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1984
Bos taurus
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Palmer, R.A.; Moss, D.S.; Haneef, I.; Borkakoti, N.
An X-ray refinement study on the binding of ribonuclease-A to cytidine-N(3)-oxide 2'-phosphate
Biochim. Biophys. Acta
785
81-88
1984
Bos taurus
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Ahmad, F.
Free energy changes in ribonuclease A denaturation. Effect of urea, guanidine hydrochloride, and lithium salts
J. Biol. Chem.
258
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1983
Bos taurus
Sagar, A.J.; Pandit, M.W.
Denaturation studies on bovine pancreatic ribonuclease. Effect of trichloroacetic acid
Biochim. Biophys. Acta
743
303-309
1983
Bos taurus
Lin, L.N.; Brandts, J.F.
Mechanism for the unfolding and refolding of ribonuclease A. Simulations using a simple model with no structural intermediates
Biochemistry
22
573-580
1983
Bos taurus
Ahmad, F.; Bigelow, C.C.
The denaturation of ribonuclease A by combinations of urea and salt denaturants
J. Mol. Biol.
131
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1979
Bos taurus
Matheson, R.R.; Scheraga, H.A.
Steps in the pathway of the thermal unfolding of ribonuclease A. A nonspecific photochemical surface-labeling study
Biochemistry
18
2437-2445
1979
Bos taurus
Leone, E.; D'Alessio, G.
Structure and properties of seminal ribonuclease
Biochem. Soc. Trans.
5
466-470
1977
Bos taurus
Weickmann, J.L.; Elson, M.; Glitz, D.G.
Purification and characterization of human pancreatic ribonuclease
Biochemistry
20
1272-1278
1981
Bos taurus, Homo sapiens
Moore, S.; Stein, W.H.
Chemical structures of pancreatic ribonuclease and deoxyribonuclease
Science
180
458-464
1973
Bos taurus
Beintema, J.J.; Campagne, R.N.; Gruber, M.
Rat pancreatic ribonuclease. I. Isolation and properties
Biochim. Biophys. Acta
310
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1973
Bos taurus, Rattus norvegicus
Pittz, E.P.; Bello, J.
Studies on bovine pancreatic ribonuclease A and model compounds in aqueous 2-methyl-2,4-pentanediol. I. Amino acid solubility, thermal reversibility of ribonuclease A, and preferential hydration of ribonuclease A crystals
Arch. Biochem. Biophys.
146
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1971
Bos taurus
Gerlsma, S.Y.; Stuur, E.R.
The effects from combining urea and an alcohol on the heat-induced reversible denaturation of ribonuclease
Int. J. Pept. Protein Res.
8
3-12
1976
Bos taurus
Bond, M.D.; Vallee, B.L.
Replacement of residues 8-22 of angiogenin with 7-21 of RNase A selectively affects protein synthesis inhibition and angiogenesis
Biochemistry
29
3341-3349
1990
Bos taurus
Hofsteenge, J.; Servis, C.; Stone, S.R.
Studies on the interaction of ribonuclease inhibitor with pancreatic ribonuclease involving differential labeling of cysteinyl residues
J. Biol. Chem.
266
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1991
Bos taurus
Neumann, U.; Hofsteenge, J.
Interaction of semisynthetic variants of RNase A with ribonuclease inhibitor
Protein Sci.
3
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1994
Bos taurus
Boix, E.; Wu, Y.; Vasandani, V.M.; Saxena, S.K.; Ardelt, W.; Ladner, J.; Youle, R.J.
Role of the N terminus in RNase A homologues: differences in catalytic activity, ribonuclease inhibitor interaction and cytotoxicity
J. Mol. Biol.
257
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1996
Bos taurus, Homo sapiens, Lithobates pipiens
Balakrishnan, R.; Ramasubbu, N.; Varughese, K.I.; Parthasarathy, R.
Crystal structures of the copper and nickel complexes of RNase A: metal-induced interprotein interactions and identification of a novel copper binding motif
Proc. Natl. Acad. Sci. USA
94
9620-9625
1997
Bos taurus
Iwaoka, M.; Juminaga, D.; Scheraga, H.A.
Regeneration of three-disulfide mutants of bovine pancreatic ribonuclease A missing the 65-72 disulfide bond: characterization of a minor folding pathway of ribonuclease A and kinetic roles of Cys65 and Cys72
Biochemistry
37
4490-4501
1998
Bos taurus
Pearson, M.A.; Karplus, P.A.; Dodge, R.W.; Laity, J.H.; Scheraga, H.A.
Crystal structures of two mutants that have implications for the folding of bovine pancreatic ribonuclease A
Protein Sci.
7
1255-1258
1998
Bos taurus
Liu, Y.; Hart, P.J.; Schlunegger, M.P.; Eisenberg, D.
The crystal structure of a 3D domain-swapped dimer of RNase A at a 2.1-A resolution
Proc. Natl. Acad. Sci. USA
95
3437-3442
1998
Bos taurus
Korn, K.; Foerster, H.H.; Hahn, U.
Phage display of RNase A and an improved method for purification of phages displaying RNases
Biol. Chem.
381
179-181
2000
Bos taurus
Fujii, T.; Doi, Y.; Ueno, H.; Hayashi, R.
Effect of mutagenic replacement of the carboxyl terminal amino acid, val124, on the properties and regeneration of bovine pancreatic ribonuclease A
J. Biochem.
127
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2000
Bos taurus
Berisio, R.; Sica, F.; Lamzin, V.S.; Wilson, K.S.; Zagari, A.; Mazzarella, L.
Atomic resolution structures of ribonuclease A at six pH values
Acta Crystallogr. Sect. D
D58
441-450
2002
Bos taurus (P61823), Bos taurus
Kadonosono, T.; Chatani, E.; Hayashi, R.; Moriyama, H.; Ueki, T.
Minimization of cavity size ensures protein stability and folding: structures of Phe46-replaced bovine pancreatic RNase A
Biochemistry
42
10651-10658
2003
Bos taurus (P61823), Bos taurus
Chatani, E.; Tanimizu, N.; Ueno, H.; Hayashi, R.
Expression of soluble bovine pancreatic ribonuclease A in Pichia pastoris and its purification and characterization
Biosci. Biotechnol. Biochem.
64
2437-2444
2000
Bos taurus
Sorrentino, S.; Barone, R.; Bucci, E.; Gotte, G.; Russo, N.; Libonati, M.; D'Alessio, G.
The two dimeric forms of RNase A
FEBS Lett.
466
35-39
2000
Bos taurus
Shin, H.C.; Song, M.C.; Scheraga, H.A.
Effect of protein disulfide isomerase on the rate-determining steps of the folding of bovine pancreatic ribonuclease A
FEBS Lett.
521
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2002
Bos taurus
Chatani, E.; Tanimizu, N.; Ueno, H.; Hayashi, R.
Structural and functional changes in bovine pancreatic ribonuclease A by the replacement of Phe120 with other hydrophobic residues
J. Biochem.
129
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2001
Bos taurus
Fujii, T.; Ueno, H.; Hayashi, R.
Significance of the four carboxyl terminal amino acid residues of bovine pancreatic ribonuclease A for structural folding
J. Biochem.
131
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2002
Bos taurus (P61823), Bos taurus
Klink, T.A.; Raines, R.T.
Conformational stability is a determinant of ribonuclease A cytotoxicity
J. Biol. Chem.
275
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2000
Bos taurus (P61823), Bos taurus
Chatani, E.; Hayashi, R.
Functional and structural roles of constituent amino acid residues of bovine pancreatic ribonuclease A
J. Biosci. Bioeng.
92
98-107
2001
Bos taurus (P61823), Bos taurus
Tanimizu, N.; Ueno, H.; Hayashi, R.
Replacement of His12 or His119 of bovine pancreatic ribonuclease A with acidic amino acid residues for the modification of activity and stability
J. Biosci. Bioeng.
94
39-44
2002
Bos taurus
Safarian, S.; Moosavi-Movahedi, A.A.
Binding patterns and kinetics of RNase A interaction with RNA
J. Protein Chem.
19
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2000
Bos taurus
Chatani, E.; Hayashi, R.; Moriyama, H.; Ueki, T.
Conformational strictness required for maximum activity and stability of bovine pancreatic ribonuclease A as revealed by crystallographic study of three Phe120 mutants at 1.4 A resolution
Protein Sci.
11
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2002
Bos taurus (P61823), Bos taurus
Ghosh, K.S.; Maiti, T.K.; Dasgupta, S.
Green tea polyphenols as inhibitors of ribonuclease A
Biochem. Biophys. Res. Commun.
325
807-811
2004
Bos taurus
Libonati, M.; Gotte, G.
Oligomerization of bovine ribonuclease A: structural and functional features of its multimers
Biochem. J.
380
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2004
Bos taurus
Gotte, G.; Laurents, D.V.; Libonati, M.
Three-dimensional domain-swapped oligomers of ribonuclease A: identification of a fifth tetramer, pentamers and hexamers, and detection of trace heptameric, octameric and nonameric species
Biochim. Biophys. Acta
1764
44-54
2006
Bos taurus (P61823), Bos taurus
Maiti, T.K.; Soumya, D.; Dasgupta, S.; Pathak, T.
3'-N-Alkylamino-3'-deoxy-ara-uridines: a new class of potential inhibitors of ribonuclease A and angiogenin
Bioorg. Med. Chem.
14
1221-1228
2006
Bos taurus (P61823)
Bucci, E.; Vitagliano, L.; Barone, R.; Sorrentino, S.; D'Alessio, G.; Graziano, G.
On the thermal stability of the two dimeric forms of ribonuclease A
Biophys. Chem.
116
89-95
2005
Bos taurus (P61823)
Yan, Y.B.; Zhang, J.; He, H.W.; Zhou, H.M.
Oligomerization and aggregation of bovine pancreatic ribonuclease A: characteristic events observed by FTIR spectroscopy
Biophys. J.
90
2525-2533
2006
Bos taurus
Kditz, J.; Ulbrich-Hofmann, R.; Arnold, U.
Probing the unfolding region of ribonuclease A by site-directed mutagenesis
Eur. J. Biochem.
271
4147-4156
2004
Bos taurus (P61823)
Mehta, R.; Kundu, A.; Kishore, N.
4-Chlorobutanol induces unusual reversible and irreversible thermal unfolding of ribonuclease A: thermodynamic, kinetic, and conformational characterization
Int. J. Biol. Macromol.
34
13-20
2004
Bos taurus
Smith, B.D.; Soellner, M.B.; Raines, R.T.
Potent inhibition of ribonuclease A by oligo(vinylsulfonic acid)
J. Biol. Chem.
278
20934-20938
2003
Bos taurus (P61823)
Sica, F.; Di Fiore, A.; Merlino, A.; Mazzarella, L.
Structure and stability of the non-covalent swapped dimer of bovine seminal ribonuclease: an enzyme tailored to evade ribonuclease protein inhibitor
J. Biol. Chem.
279
36753-36760
2004
Bos taurus
Safarian, S.; Moosavi-Movahedi, A.A.; Hosseinkhani, S.; Xia, Z.; Habibi-Rezaei, M.; Hosseini, G.; Sorenson, C.; Sheibani, N.
The structural and functional studies of His119 and His12 in RNase A via chemical modification
J. Protein Chem.
22
643-654
2003
Bos taurus
Schultz, D.A.; Friedman, A.M.; White, M.A.; Fox, R.O.
The crystal structure of the cis-proline to glycine variant (P114G) of ribonuclease A
Protein Sci.
14
2862-2870
2005
Bos taurus (P61823)
Larson, S.B.; Day, J.S.; Cudney, R.; McPherson, A.
A new crystal form of bovine pancreatic RNase A in complex with 2-deoxyguanosine-5-monophosphate
Acta Crystallogr. Sect. F
F63
728-733
2007
Bos taurus
Tan, C.J.; Tong, Y.W.
Preparation of superparamagnetic ribonuclease A surface-imprinted submicrometer particles for protein recognition in aqueous media
Anal. Chem.
79
299-306
2007
Bos taurus
Yakovlev, G.I.; Mitkevich, V.A.; Struminskaya, N.K.; Varlamov, V.P.; Makarov, A.A.
Low molecular weight chitosan is an efficient inhibitor of ribonucleases
Biochem. Biophys. Res. Commun.
357
584-588
2007
Bos taurus
Merkley, E.D.; Bernard, B.; Daggett, V.
Conformational changes below the Tm: molecular dynamics studies of the thermal pretransition of ribonuclease A
Biochemistry
47
880-892
2008
Bos taurus (P61823), Bos taurus
Wang, S.; Li, H.
Radical scavenging activity of ribonuclease inhibitor from cow placenta
Biochemistry (Moscow)
71
520-524
2006
Bos taurus
Torrent, J.; Font, J.; Herberhold, H.; Marchal, S.; Ribo, M.; Ruan, K.; Winter, R.; Vilanova, M.; Lange, R.
The use of pressure-jump relaxation kinetics to study protein folding landscapes
Biochim. Biophys. Acta
1764
489-496
2006
Bos taurus
Pouckova, P.; Morbio, M.; Vottariello, F.; Laurents, D.V.; Matousek, J.; Soucek, J.; Gotte, G.; Donadelli, M.; Costanzo, C.; Libonati, M.
Cytotoxicity of polyspermine-ribonuclease A and polyspermine-dimeric ribonuclease A
Bioconjug. Chem.
18
1946-1955
2007
Bos taurus
Font, J.; Benito, A.; Torrent, J.; Lange, R.; Ribo, M.; Vilanova, M.
Pressure- and temperature-induced unfolding studies: thermodynamics of core hydrophobicity and packing of ribonuclease A
Biol. Chem.
387
285-296
2006
Bos taurus (P61823)
Leonidas, D.D.; Maiti, T.K.; Samanta, A.; Dasgupta, S.; Pathak, T.; Zographos, S.E.; Oikonomakos, N.G.
The binding of 3-N-piperidine-4-carboxyl-3-deoxy-ara-uridine to ribonuclease A in the crystal
Bioorg. Med. Chem.
14
6055-6064
2006
Bos taurus (P61823)
Font, J.; Torrent, J.; Ribo, M.; Laurents, D.V.; Balny, C.; Vilanova, M.; Lange, R.
Pressure-jump-induced kinetics reveals a hydration dependent folding/unfolding mechanism of ribonuclease A
Biophys. J.
91
2264-2274
2006
Bos taurus
Polydoridis, S.; Leonidas, D.D.; Oikonomakos, N.G.; Archontis, G.
Recognition of ribonuclease a by 3-5-pyrophosphate-linked dinucleotide inhibitors: a molecular dynamics/continuum electrostatics analysis
Biophys. J.
92
1659-1672
2007
Bos taurus (P61823)
Noronha, M.; Lima, J.C.; Paci, E.; Santos, H.; Macanita, A.L.
Tracking local conformational changes of ribonuclease A using picosecond time-resolved fluorescence of the six tyrosine residues
Biophys. J.
92
4401-4414
2007
Bos taurus
Choi, Y.; Lee, J.H.; Hwang, S.; Kim, J.K.; Jeong, K.; Jung, S.
Retardation of the unfolding process by single N-glycosylation of ribonuclease A based on molecular dynamics simulations
Biopolymers
89
114-123
2008
Bos taurus (P61823), Bos taurus
Cozza, G.; Moro, S.; Gotte, G.
Elucidation of the ribonuclease A aggregation process mediated by 3D domain swapping: a computational approach reveals possible new multimeric structures
Biopolymers
89
26-39
2008
Bos taurus (P61823), Bos taurus
Hsu, C.Y.; Lin, H.Y.; Thomas, J.L.; Wu, B.T.; Chou, T.C.
Incorporation of styrene enhances recognition of ribonuclease A by molecularly imprinted polymers
Biosens. Bioelectron.
22
355-363
2006
Bos taurus
Johnson, R.J.; Lin, S.R.; Raines, R.T.
A ribonuclease zymogen activated by the NS3 protease of the hepatitis C virus
FEBS J.
273
5457-5465
2006
Bos taurus, Homo sapiens
Pradeep, L.; Shin, H.C.; Scheraga, H.A.
Correlation of folding kinetics with the number and isomerization states of prolines in three homologous proteins of the RNase family
FEBS Lett.
580
5029-5032
2006
Bos taurus, Lithobates pipiens
Volynskaya, A.V.; Kasumov, E.A.; Goldanskii, V.I.
An evidence for the equilibrium unfolding intermediates of ribonuclease A by tritium labeling method
Int. J. Biol. Macromol.
39
256-264
2006
Bos taurus
Li, H.; Wang, S.
Kinetics of inhibition of ribonuclease A by Pholiota Nameko polysaccharide
Int. J. Biol. Macromol.
40
134-138
2007
Bos taurus
Leich, F.; Koeditz, J.; Ulbrich-Hofman, R.; Arnold, U.
Tandemization endows bovine pancreatic ribonuclease with cytotoxic activity
J. Mol. Biol.
358
1305-1313
2006
Bos taurus (P61823), Bos taurus
Anissimova, M.V.; Baek, W.O.; Varlamov, V.P.; Mrabet, N.T.; Vijayalakshmi, M.A.
Natural and chemically induced oligomeric ribonucleases: structural study by immobilized metal ion affinity electrophoresis and their functional relationship
J. Mol. Recognit.
19
287-298
2006
Bos taurus
Sardar, P.S.; Maity, S.S.; Ghosh, S.; Chatterjee, J.; Maiti, T.K.; Dasgupta, S.
Characterization of the tryptophan residues of human placental ribonuclease inhibitor and its complex with bovine pancreatic ribonuclease A by steady-state and time-resolved emission spectroscopy
J. Phys. Chem. B
110
21349-21356
2006
Bos taurus
Tan, C.J.; Tong, Y.W.
The effect of protein structural conformation on nanoparticle molecular imprinting of ribonuclease A using miniemulsion polymerization
Langmuir
23
2722-2730
2007
Bos taurus
Skewis, L.R.; Reinhard, B.M.
Spermidine modulated ribonuclease activity probed by RNA plasmon rulers
Nano Lett.
8
214-220
2008
Bos taurus
Pouckova, P.; Skvor, J.; Gotte, G.; Vottariello, F.; Slavik, J.T.; Matousek, J.; Laurents, D.V.; Libonati, M.; Soucek, J.
Some biological actions of PEG-conjugated RNase A oligomers
Neoplasma
53
79-85
2006
Bos taurus
Schnell, C.; Scharnagl, C.; Friedrich, J.
Hole burning spectroscopy of ribonuclease A
Phys. Chem. Chem. Phys.
8
1315-1320
2006
Bos taurus
Trifonova, E.A.; Sapotsky, M.V.; Komarova, M.L.; Scherban, A.B.; Shumny, V.K.; Polyakova, A.M.; Lapshina, L.A.; Kochetov, A.V.; Malinovsky, V.I.
Protection of transgenic tobacco plants expressing bovine pancreatic ribonuclease against tobacco mosaic virus
Plant Cell Rep.
26
1121-1126
2007
Bos taurus (P61823), Bos taurus
Moosavi-Movahedi, A.A.; Gharanfoli, M.; Jalili, S.; Ahmad, F.; Chamani, J.; Hakimelahi, G.H.; Sadeghi, M.; Amani, M.; Saboury, A.A.
The correlation of RNase A enzymatic activity with the changes in the distance between Nepsilon2-His12 and N delta1-His119 upon addition of stabilizing and destabilizing salts
Protein J.
25
117-125
2006
Bos taurus (P61823)
Zhang, J.; He, H.W.; Wang, Q.; Yan, Y.B.
Sequential events in ribonuclease A thermal unfolding characterized by two-dimensional infrared correlation spectroscopy
Protein Pept. Lett.
13
33-40
2006
Bos taurus
Younus, H.; Ulbrich-Hofmann, R.; Saleemuddin, M.
Inhibition of pancreatic ribonuclease A aggregation by antibodies raised against the native enzyme and its N-terminal dodecapeptide
Protein Pept. Lett.
13
673-677
2006
Bos taurus
Font, J.; Benito, A.; Lange, R.; Ribo, M.; Vilanova, M.
The contribution of the residues from the main hydrophobic core of ribonuclease A to its pressure-folding transition state
Protein Sci.
15
1000-1009
2006
Bos taurus
Moussaoui, M.; Cuchillo, C.M.; Nogues, M.V.
A phosphate-binding subsite in bovine pancreatic ribonuclease A can be converted into a very efficient catalytic site
Protein Sci.
16
99-109
2007
Bos taurus (P61823), Bos taurus
Simons, B.L.; Kaplan, H.; Fournier, S.M.; Cyr, T.; Hefford, M.A.
A novel cross-linked RNase A dimer with enhanced enzymatic properties
Proteins
66
183-195
2007
Bos taurus
Ghosh, K.S.; Maiti, T.K.; Debnath, J.; Dasgupta, S.
Inhibition of ribonuclease A by polyphenols present in green tea
Proteins
69
566-580
2007
Bos taurus (P61823)
Pradeep, L.; Kurinov, I.; Ealick, S.E.; Scheraga, H.A.
Implementation of a k/k(0) method to identify long-range structure in transition states during conformational folding/unfolding of proteins
Structure
15
1178-1189
2007
Bos taurus (P61823), Bos taurus
Wang, L.; Wu, Y.; Meersman, F.
Clarification of the thermally-induced pretransition of ribonuclease A in solution by principal component analysis and two-dimensional correlation infrared spectroscopy
Vib. Spectrosc.
42
201-205
2006
Bos taurus
-
Gautschi, M.; Beintema, J.J.
Selection against glycosylation in ruminant pancreatic ribonucleases by replacements in the ancestral carbohydrate attachment site
Biochem. Genet.
46
446-450
2008
Bos taurus
Merlino, A.; Krauss, I.R.; Perillo, M.; Mattia, C.A.; Ercole, C.; Picone, D.; Vergara, A.; Sica, F.
Towards an antitumor form of bovine pancreatic ribonuclease: The crystal structure of three non-covalent dimeric mutants
Biopolymers
91
1029-1037
2009
Bos taurus (P61823), Bos taurus
Yagi, D.; Yamada, T.; Kurihara, K.; Ohnishi, Y.; Yamashita, M.; Tamada, T.; Tanaka, I.; Kuroki, R.; Niimura, N.
A neutron crystallographic analysis of phosphate-free ribonuclease A at 1.7 A resolution
Acta Crystallogr. Sect. D
65
892-899
2009
Bos taurus (P61823), Bos taurus
Tsirkone, V.G.; Dossi, K.; Drakou, C.; Zographos, S.E.; Kontou, M.; Leonidas, D.D.
Inhibitor design for ribonuclease A: the binding of two 5-phosphate uridine analogues
Acta Crystallogr. Sect. F
65
671-677
2009
Bos taurus (P61823)
Larson, S.B.; Day, J.S.; Nguyen, C.; Cudney, R.; McPherson, A.
Structure of bovine pancreatic ribonuclease complexed with uridine 5-monophosphate at 1.60 A resolution
Acta Crystallogr. Sect. F
66
113-120
2010
Bos taurus (P61823), Bos taurus
Allgaier, S.; Weiland, N.; Hamad, I.; Kempken, F.
Expression of ribonuclease A and ribonuclease N1 in the filamentous fungus Neurospora crassa
Appl. Microbiol. Biotechnol.
85
1041-1049
2010
Bos taurus (P61823)
Pearce, F.G.; Griffin, M.D.; Gerrard, J.A.
Does domain swapping improve the stability of RNase A?
Biochem. Biophys. Res. Commun.
382
114-118
2009
Bos taurus
Ramadan, D.; Rancy, P.C.; Nagarkar, R.P.; Schneider, J.P.; Thorpe, C.
Arsenic(III) species inhibit oxidative protein folding in vitro
Biochemistry
48
424-432
2009
Bos taurus
Dickson, K.A.; Raines, R.T.
Silencing an inhibitor unleashes a cytotoxic enzyme
Biochemistry
48
5051-5053
2009
Bos taurus
Almarza, J.; Rincon, L.; Bahsas, A.; Brito, F.
Molecular mechanism for the denaturation of proteins by urea
Biochemistry
48
7608-7613
2009
Bos taurus
Debnath, J.; Dasgupta, S.; Pathak, T.
Nucleoside-amino acid conjugates: An alternative route to the design of ribonuclease A inhibitors
Bioorg. Med. Chem.
17
4921-4927
2009
Bos taurus (P61823)
Debnath, J.; Dasgupta, S.; Pathak, T.
Inhibition of ribonuclease A by nucleoside-dibasic acid conjugates
Bioorg. Med. Chem.
17
6491-6495
2009
Bos taurus (P61823)
Pecher, P.; Arnold, U.
The effect of additional disulfide bonds on the stability and folding of ribonuclease A
Biophys. Chem.
141
21-28
2009
Bos taurus (P61823)
Ercole, C.; Colamarino, R.A.; Pizzo, E.; Fogolari, F.; Spadaccini, R.; Picone, D.
Comparison of the structural and functional properties of RNase A and BS-RNase: a stepwise mutagenesis approach
Biopolymers
91
1009-1017
2009
Bos taurus (P00669), Bos taurus
Vila, R.; Benito, A.; Ribo, M.; Vilanova, M.
Mapping the stability clusters in bovine pancreatic ribonuclease A
Biopolymers
91
1038-1047
2009
Bos taurus (P61823), Bos taurus
Holloway, D.E.; Chavali, G.B.; Leonidas, D.D.; Baker, M.D.; Acharya, K.R.
Influence of naturally-occurring 5-pyrophosphate-linked substituents on the binding of adenylic inhibitors to ribonuclease a: an X-ray crystallographic study
Biopolymers
91
995-1008
2009
Bos taurus (P61823)
Lee, D.H.; Kim, S.G.; Kweon, D.H.; Seo, J.H.
Folding machineries displayed on a cation-exchanger for the concerted refolding of cysteine- or proline-rich proteins
BMC Biotechnol.
9
27
2009
Bos taurus
Lopez-Alonso, J.P.; Bruix, M.; Font, J.; Ribo, M.; Vilanova, M.; Jimenez, M.A.; Santoro, J.; Gonzalez, C.; Laurents, D.V.
NMR spectroscopy reveals that RNase A is chiefly denatured in 40% acetic acid: implications for oligomer formation by 3D domain swapping
J. Am. Chem. Soc.
132
1621-1630
2010
Bos taurus
Day, I.J.; Maeda, K.; Paisley, H.J.; Mok, K.H.; Hore, P.J.
Refolding of ribonuclease A monitored by real-time photo-CIDNP NMR spectroscopy
J. Biomol. NMR
44
77-86
2009
Bos taurus (P61823), Bos taurus
Samanta, A.; Leonidas, D.D.; Dasgupta, S.; Pathak, T.; Zographos, S.E.; Oikonomakos, N.G.
Morpholino, piperidino, and pyrrolidino derivatives of pyrimidine nucleosides as inhibitors of ribonuclease A: synthesis, biochemical, and crystallographic evaluation
J. Med. Chem.
52
932-942
2009
Bos taurus (P61823)
Shahhoseini, M.; Rabbani Chadegani, A.; Abdosamadi, S.
Evidence for the structural stability of ribonucleoprotein LMG(160) under ribonuclease-A treatment
Mol. Cell. Biochem.
321
65-72
2009
Bos taurus
Goldschmidt, L.; Teng, P.K.; Riek, R.; Eisenberg, D.
Identifying the amylome, proteins capable of forming amyloid-like fibrils
Proc. Natl. Acad. Sci. USA
107
3487-3492
2010
Bos taurus
Dechene, M.; Wink, G.; Smith, M.; Swartz, P.; Mattos, C.
Multiple solvent crystal structures of ribonuclease A: an assessment of the method
Proteins
76
861-881
2009
Bos taurus (P61823), Bos taurus
Kurpiewska, K.; Font, J.; Ribo, M.; Vilanova, M.; Lewi?ski, K.
X-ray crystallographic studies of RNase A variants engineered at the most destabilizing positions of the main hydrophobic core: further insight into protein stability
Proteins
77
658-669
2009
Bos taurus (P61823), Bos taurus
Ghosh, U.; Giri, K.; Bhattacharyya, N.P.
Interaction of aurintricarboxylic acid (ATA) with four nucleic acid binding proteins DNase I, RNase A, reverse transcriptase and Taq polymerase
Spectrochim. Acta A. Mol. Biomol. Spectrosc.
74
1145-1151
2009
Bos taurus
Arai, K.; Kumakura, F.; Iwaoka, M.
Characterization of kinetic and thermodynamic phases in the prefolding process of bovine pancreatic ribonuclease A coupled with fast SS formation and SS reshuffling
Biochemistry
49
10535-10542
2010
Bos taurus
Doucet, N.; Khirich, G.; Kovrigin, E.L.; Loria, J.P.
Alteration of hydrogen bonding in the vicinity of histidine 48 disrupts millisecond motions in RNase A
Biochemistry
50
1723-1730
2011
Bos taurus
Cuchillo, C.M.; Nogues, M.V.; Raines, R.T.
Bovine pancreatic ribonuclease: fifty years of the first enzymatic reaction mechanism
Biochemistry
50
7835-7841
2011
Bos taurus
Vottariello, F.; Costanzo, C.; Gotte, G.; Libonati, M.
Zero-length dimers of ribonuclease A: further characterization and no evidence of cytotoxicity
Bioconjug. Chem.
21
635-645
2010
Bos taurus
Debnath, J.; Dasgupta, S.; Pathak, T.
Comparative inhibitory activity of 3- and 5-functionalized nucleosides on ribonuclease A
Bioorg. Med. Chem.
18
8257-8263
2010
Bos taurus
Samanta, A.; Dasgupta, S.; Pathak, T.
5'-modified pyrimidine nucleosides as inhibitors of ribonuclease A
Bioorg. Med. Chem.
19
2478-2484
2011
Bos taurus (P61823)
Arnold, U.; Leich, F.; Neumann, P.; Lilie, H.; Ulbrich-Hofmann, R.
Crystal structure of RNase A tandem enzymes and their interaction with the cytosolic ribonuclease inhibitor
FEBS J.
278
331-340
2011
Bos taurus (P61823)
Geiger, R.; Gautschi, M.; Thor, F.; Hayer, A.; Helenius, A.
Folding, quality control, and secretion of pancreatic ribonuclease in live cells
J. Biol. Chem.
286
5813-5822
2011
Bos taurus, Homo sapiens
Miller, K.H.; Karr, J.R.; Marqusee, S.
A hinge region cis-proline in ribonuclease A acts as a conformational gatekeeper for C-terminal domain swapping
J. Mol. Biol.
400
567-578
2010
Bos taurus
Graham, D.; Greminger, J.
On the information expressed in enzyme structure: more lessons from ribonuclease A
Mol. Divers.
15
769-779
2011
Bos taurus
Miller, K.H.; Marqusee, S.
Propensity for C-terminal domain swapping correlates with increased regional flexibility in the C-terminus of RNase A
Protein Sci.
20
1735-1744
2011
Bos taurus
Doucet, N.; Jayasundera, T.; Simonovic, M.; Loria, J.
The crystal structure of ribonuclease a in complex with thymidine-3'-monophosphate provides further insight into ligand binding
Proteins
78
2459-2468
2010
Bos taurus (P61823), Bos taurus
Fiorini, C.; Gotte, G.; Donnarumma, F.; Picone, D.; Donadelli, M.
Bovine seminal ribonuclease triggers Beclin1-mediated autophagic cell death in pancreatic cancer cells
Biochim. Biophys. Acta
1843
976-984
2014
Bos taurus
Merlino, A.; Picone, D.; Ercole, C.; Balsamo, A.; Sica, F.
Chain termini cross-talk in the swapping process of bovine pancreatic ribonuclease
Biochimie
94
1108-1118
2012
Bos taurus (P61823), Bos taurus
Tomita, S.; Nagasaki, Y.; Shiraki, K.
Different mechanisms of action of poly(ethylene glycol) and arginine on thermal inactivation of lysozyme and ribonuclease A
Biotechnol. Bioeng.
109
2543-2552
2012
Bos taurus
Gagne, D.; Doucet, N.
Structural and functional importance of local and global conformational fluctuations in the RNase A superfamily
FEBS J.
280
5596-5607
2013
Gallus gallus, Homo sapiens, Danio rerio (A5HAK0), Bos taurus (P00669), Rattus norvegicus (P00684), Lithobates pipiens (P22069)
Amiri, R.; Bordbar, A.K.; Laurents, D.V.; Khosropour, A.R.; Mohammadpoor-Baltork, I.
Thermal stability and enzymatic activity of RNase A in the presence of cationic gemini surfactants
Int. J. Biol. Macromol.
50
1151-1157
2012
Bos taurus
Sakaue, H.; Kinouchi, T.; Fujii, N.; Fujii, N.; Takata, T.
Isomeric replacement of a single aspartic acid induces a marked change in protein function the example of ribonuclease A
ACS Omega
2
260-267
2017
Bos taurus (P61823)
-
Vermeire, K.; Allan, S.; Provinciael, B.; Hartmann, E.; Kalies, K.U.
Ribonuclease-neutralized pancreatic microsomal membranes from livestock for in vitro co-translational protein translocation
Anal. Biochem.
484
102-104
2015
Sus scrofa (P00671), Bos taurus (P61823), Ovis aries (P67927)
Kurpiewska, K.; Dziubek, K.; Katrusiak, A.; Font, J.; Ribo, M.; Vilanova, M.; Lewinski, K.
Structural investigation of ribonuclease A conformational preferences using high pressure protein crystallography
Chem. Phys.
468
53-62
2016
Bos taurus (P61823)
-
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