Information on EC 3.4.23.21 - Rhizopuspepsin

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The expected taxonomic range for this enzyme is: Rhizopus

EC NUMBER
COMMENTARY hide
3.4.23.21
-
RECOMMENDED NAME
GeneOntology No.
Rhizopuspepsin
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
hydrolysis of proteins with broad specificity similar to that of pepsin A, preferring hydrophobic residues at P1 and P1'. Clots milk and activates trypsinogen. Does not cleave Gln4-His, but does cleave His10-/-Leu and Val12-/-Glu in B chain of insulin
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of peptide bond
-
-
aspartic endopeptidase
-
CAS REGISTRY NUMBER
COMMENTARY hide
9074-09-3
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SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
Ac-Ala-Ala-(4-nitro)Phe-Ala-Ala + H2O
?
show the reaction diagram
-
-
-
-
-
casein + H2O
?
show the reaction diagram
FVNQHLCGSHLVEALYLVCGERGFFYTPKA + H2O
L-Phe + VNQH + LCGSH + L-Leu + L-Val + L-Glu + L-Ala + L-Leu + L-Tyr + LVCG + ERG + L-Phe + L-Phe + YTPKA
show the reaction diagram
KAIEF p-nitrophenylalanine-RL + H2O
KAIEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KLIEF p-nitrophenylalanine-RL + H2O
KLIEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPAEF p-nitrophenylalanine-RL + H2O
KPAEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPALF p-nitrophenylalanine-RL + H2O
KPALF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPDEF p-nitrophenylalanine-RL + H2O
KPDEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPIAF p-nitrophenylalanine-RL + H2O
KPIAF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPIDF p-nitrophenylalanine-RL + H2O
KPIDF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPIEF p-nitrophenylalanine-RL + H2O
KPIEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPILF p-nitrophenylalanine-RL + H2O
KPILF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPISF p-nitrophenylalanine-RL + H2O
KPISF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPLEF p-nitrophenylalanine-RL + H2O
KPLEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPREF p-nitrophenylalanine-RL + H2O
KPREF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPRRPYILKRGSYYY + H2O
KPRRPYIL + KRGSYYY
show the reaction diagram
-
synthetic neurotensin-like peptide, cleavage site specificity
-
-
?
KPSEF p-nitrophenylalanine-RL + H2O
KPSEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
KPVSY(4-NO2-F)RL + H2O
?
show the reaction diagram
-
-
-
-
?
KPVSY(NO2)FRL + H2O
?
show the reaction diagram
-
-
-
?
KSIEF p-nitrophenylalanine-RL + H2O
KSIEF + p-nitrophenylalanine-RL
show the reaction diagram
-
-
-
?
L-Orn-L-Leu-D-Phe-L-Pro-L-Val-OH + H2O
L-Orn + L-Val + L-Leu-D-Phe-L-Pro-OH
show the reaction diagram
-
open chain of gramicidin S that is prepared by the treatment of gramicidin S with Bacillus subtilis alkaline protease
-
-
Lys-Pro-Ala-Lys-Phe-(NO2)Phe-Arg-Leu + H2O
Lys-Pro-Ala-Lys-Phe + (NO2)Phe-Arg-Leu
show the reaction diagram
-
-
-
?
Lys-Pro-Ile-Glu-Phe-(NO2)Phe-Arg-Leu + H2O
Lys-Pro-Ile-Glu-Phe + (NO2)Phe-Arg-Leu
show the reaction diagram
-
-
-
?
Na-Polyglutamate + H2O
Glu + Glu-Glu + unknown compound
show the reaction diagram
-
-
-
-
oxidized insulin B chain + H2O
?
show the reaction diagram
-
-
-
?
Oxidized insulin B-chain + H2O
Proteolytically cleaved oxidized insulin B-chain
show the reaction diagram
Peptides + H2O
?
show the reaction diagram
Polylysine-HCl + H2O
Lys-Lys + Lys-Lys-Lys + higher oligolysine
show the reaction diagram
-
-
-
-
Z-Ala-Ala-Lys-Ala-Ala-Ala + H2O
Z-Ala-Ala-Lys + Ala-Ala-Ala
show the reaction diagram
-
-
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Cu2+
-
1 mM, activates
Mn2+
-
1 mM, activates
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1,2-epoxy-3-(4-nitrophenoxy)propane
Ac-NA-Nva-3-hydroxy 4-amino 6-methyl heptanoic acid-Nva-NA-amide
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-
Ac-NA-Nva-3-hydroxy 4-amino 6-methyl heptanoyl-Nva-NA-amide
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Ac-NA-Val-3-hydroxy 4-amino 6-methyl heptanoic acid-2-aminobutanoyl-NA-amide
-
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Ac-NA-Val-3-hydroxy 4-amino 6-methyl heptanoyl-Abu-NA-amide
-
-
acetylpepstatin
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Streptomyces pepsin inhibitor
Diazoacetyl-DL-norleucine methyl ester
Pepstatin
pepstatin A
-
-
Peptide inhibitor
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sequence: D-His-Pro-Phe-His-Phe(Psi)[CH2-NH]Phe-Val-Tyr
-
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.23 - 1.1
Ac-Ala-Ala-Lys-(4-nitro)Phe-Ala-Ala
0.006 - 0.269
KAIEF p-nitrophenylalanine-RL
0.007 - 0.139
KLIEF p-nitrophenylalanine-RL
0.008 - 0.089
KPAEF p-nitrophenylalanine-RL
0.004 - 0.013
KPALF p-nitrophenylalanine-RL
0.01 - 0.065
KPDEF p-nitrophenylalanine-RL
0.006 - 0.157
KPIAF p-nitrophenylalanine-RL
0.003 - 0.211
KPIDF p-nitrophenylalanine-RL
0.002 - 0.057
KPIEF p-nitrophenylalanine-RL
0.006 - 0.056
KPILF p-nitrophenylalanine-RL
0.003 - 0.047
KPISF p-nitrophenylalanine-RL
0.004 - 0.101
KPLEF p-nitrophenylalanine-RL
0.008
KPREF p-nitrophenylalanine-RL
-
pH 3.0, 37C
0.007 - 0.108
KPSEF p-nitrophenylalanine-RL
0.0034
KPVSY(4-NO2-F)RL
-
-
0.0034
KPVSY(NO2)FRL
-
pH 7.4, 37C
0.009 - 0.21
KSIEF p-nitrophenylalanine-RL
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
5 - 46
KAIEF p-nitrophenylalanine-RL
5 - 38
KLIEF p-nitrophenylalanine-RL
21 - 22
KPAEF p-nitrophenylalanine-RL
9 - 13
KPALF p-nitrophenylalanine-RL
3 - 8
KPDEF p-nitrophenylalanine-RL
3 - 17
KPIAF p-nitrophenylalanine-RL
0.77 - 6.08
KPIDF p-nitrophenylalanine-RL
6 - 41
KPIEF p-nitrophenylalanine-RL
0.23 - 3
KPILF p-nitrophenylalanine-RL
3 - 4
KPISF p-nitrophenylalanine-RL
7
KPLEF p-nitrophenylalanine-RL
Rhizopus sp.
-
pH 3.0, 37C, wild-type and chimeric enzyme
18
KPREF p-nitrophenylalanine-RL
Rhizopus sp.
-
pH 3.0, 37C
6 - 7
KPSEF p-nitrophenylalanine-RL
55
KPVSY(4-NO2-F)RL
Rhizopus microsporus var. chinensis
-
-
55
KPVSY(NO2)FRL
Rhizopus sp.
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pH 7.4, 37C
1.8 - 13
KSIEF p-nitrophenylalanine-RL
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.000077
Ac-NA-Nva-3-hydroxy 4-amino 6-methyl heptanoic acid-Nva-NA-amide
-
pH 3.0, 37C
0.0000029
Ac-NA-Val-3-hydroxy 4-amino 6-methyl heptanoic acid-2-aminobutanoyl-NA-amide
-
pH 3.0, 37C
0.0000006
pepstatin A
-
pH 3.0, 37C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2.9 - 4.5
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varying maximum dependent on the substrate
3 - 4
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trypsinogen activation
3.5 - 4
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casein
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2 - 6.5
-
-
5.5 - 7.5
-
95% of maximal activity within this range
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25
-
assay at
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
34000
-
1 * 34000
34230
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Rhizopus chinensis, rhizopuspepsin II, calculation from amino acid sequence
34340
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Rhizopus chinensis, rhizopuspepsin I, calculation from amino acid sequence
36000
-
Rhizopus hangchow, FPLC gel filtration
37600
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1 * 37600, Rhizopus hangchow, SDS-PAGE
40300
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Rhizopus niveus, low speed sedimentation without reaching maximum
additional information
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
additional information
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structure analysis
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structure at: 5.5 A resolution
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structure and refinement at 1.8 A resolution
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three-dimensional structure of the complex of rhizopuspepsin with pepstatin at 2.5 A resolution
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chimeric enzymes, containing domains of porcine pepsinogen
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crystal structure determined at pH 4.6, 7.0, and 8.0, using hanging-drop method, space group P2(1)2(1)2(1) unit-cell parameters a 0 60.48 A, b : 60.52 A, c : 106.87 A at pH 4.6, a : 60.19 A, b : 60.79 A, c : 106.99 A at pH 7.0, a : 60.37 A, b : 60.58 A, c : 107.36 A at pH 8.0, enzyme cannot be crystallized outside the pH range 4.6-8.0
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crystals with typical dimensions of 0.3 x 0.3 x 0.6 mm, orthorhombic crystals, space group P2(1)2(1)2(1), with dimensions a : 60.18A, b : 71.10A, and c : 81.21A
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pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2
-
0.1 M tartrate buffer, 24 h, loses most of its activity, retains its high activity if incubated in 0.1 M tartrate buffer of pH 3.0-4.0 or in 1.0 M acetate buffer of pH 4.5-5.5
30606
6.5 - 7.5
-
0.1 M phosphate buffer, 24 h, loses most of its activity, retains its high activity if incubated in 0.1 M tartrate buffer of pH 3.0-4.0 or in 1.0 M acetate buffer of 4.5-5.5
30606
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 45
-
purified enzyme, stable
45
-
pH 3, 5 min, stable
50
-
pH 3, 5 min, complete inactivation
58
-
10 min, 90% loss of activity
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, pure rhizopuspepsin can be stored frozen for at least 4 months
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
2 major isoenzymes with pI 5.1 and 5.8, 3 minor isoenzymes with pI 7.35, 7.41 and 7.9, partial
-
native enzyme 91fold by ammonium sulfate fractionation, ion exchange chromatography and gel filtration
-
native enzyme from culture medium
native enzyme from culture medium by ammonium sulfate precipitation and pepstatin affinity chromatography, further separation of isozymes by isoelectric focusing, recombinant enzyme from Escherichia coli inclusion bodies
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proenzyme
-
rapid, efficient 1-step purification, high-performance hydrophobic interaction chromatography of proteins on reversed-phase supports coated with non-ionic surfactants of polyoxyethylene type
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DNA sequence determination
expresssion in Escherichia coli in inclusion bodies
-
rhizopuspepsinogen gene transformed into Escherichia coli DH5-alpha
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Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
solubilization and refolding of recombinant enzyme from Escherichia coli inclusion bodies
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
medicine
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design of a selective and potent inhibitor is crucial since aspartic proteinases known to be involved in many diseases such as cancer and malaria
molecular biology
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rhizopuspepsin is a good model enzyme to investigate acid proteinases