Information on EC 6.1.1.16 - Cysteine-tRNA ligase

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The expected taxonomic range for this enzyme is: Eukaryota, Bacteria, Archaea

EC NUMBER
COMMENTARY
6.1.1.16
-
RECOMMENDED NAME
GeneOntology No.
Cysteine-tRNA ligase
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
ATP + L-cysteine + tRNACys = AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys = AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
the enzyme shows dual substrate specificity, Glu103 is critical for proline binding, Pro100 is essential for cysteine binding
-
ATP + L-cysteine + tRNACys = AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
substrate binding mechanism, induced fit
P21888
ATP + L-cysteine + tRNACys = AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
the class I tRNA synthetase CysRS is rate-limited by release of aminoacyl-tRNA, class I synthetases share two signature motifs, HIGH and KMSKS, and build their active site by the ubiquitous Rossmann nucleotide-binding fold, the tight aminoacyl-tRNA product binding by class I enzymes correlates with the ability of EF-Tu to form a ternary complex with class I but not class II synthetases, and the further capacity of this protein to enhance the rate of aminoacylation by class I synthetases
-
REACTION TYPE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Aminoacylation
-
-
-
-
Aminoacylation
-
-
Aminoacylation
-, Q9UWP2
-
Aminoacylation
-
-
Aminoacylation
Bacillus subtilis 168T
-
-
-
esterification
-
-
-
-
esterification
-
-
esterification
-, Q9UWP2
-
esterification
-
-
esterification
Bacillus subtilis 168T
-
-
-
PATHWAY
KEGG Link
MetaCyc Link
Aminoacyl-tRNA biosynthesis
-
tRNA charging
-
SYSTEMATIC NAME
IUBMB Comments
L-Cysteine:tRNACys ligase (AMP-forming)
-
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
class I CysRS
-
-
class I cysteinyl-tRNA synthetase
-
-
CysRS
-
-
-
-
CysRS
-
-
Cysteine translase
-
-
-
-
Cysteine translase
-
-
Cysteine translase
Bacillus subtilis 168T
-
-
-
Cysteine translase
-
-
Cysteine translase
-
-
Cysteine translase
Q9UWP2
-
Cysteine translase
-
-
Cysteine--tRNA ligase
-
-
-
-
Cysteine--tRNA ligase
-
-
Cysteine--tRNA ligase
-
-
Cysteine--tRNA ligase
-
-
Cysteine--tRNA ligase
Q9UWP2
-
Cysteine--tRNA ligase
-
-
Cysteinyl-transfer ribonucleate synthetase
-
-
-
-
Cysteinyl-transfer ribonucleate synthetase
-
-
Cysteinyl-transfer ribonucleate synthetase
Bacillus subtilis 168T
-
-
-
Cysteinyl-transfer ribonucleate synthetase
-
-
Cysteinyl-transfer ribonucleate synthetase
-
-
Cysteinyl-transfer ribonucleate synthetase
Q9UWP2
-
Cysteinyl-transfer ribonucleate synthetase
-
-
Cysteinyl-transfer RNA synthetase
-
-
-
-
Cysteinyl-transfer RNA synthetase
-
-
Cysteinyl-transfer RNA synthetase
Bacillus subtilis 168T
-
-
-
Cysteinyl-transfer RNA synthetase
-
-
Cysteinyl-transfer RNA synthetase
-
-
Cysteinyl-transfer RNA synthetase
Q9UWP2
-
Cysteinyl-transfer RNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
Bacillus subtilis 168T
-
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
Q9UWP2
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
Cysteinyl-tRNA synthetase
-
-
prolyl-cysteinyl-tRNA synthetase
-
-
Synthetase, cysteinyl-transfer ribonucleate
-
-
-
-
Synthetase, cysteinyl-transfer ribonucleate
-
-
Synthetase, cysteinyl-transfer ribonucleate
Bacillus subtilis 168T
-
-
-
Synthetase, cysteinyl-transfer ribonucleate
-
-
Synthetase, cysteinyl-transfer ribonucleate
-
-
Synthetase, cysteinyl-transfer ribonucleate
Q9UWP2
-
Synthetase, cysteinyl-transfer ribonucleate
-
-
CAS REGISTRY NUMBER
COMMENTARY
37318-56-2
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
strain 168T, gene cysS
-
-
Manually annotated by BRENDA team
Bacillus subtilis 168T
strain 168T, gene cysS
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
class I enzyme
Uniprot
Manually annotated by BRENDA team
K-12; temperature-sensitive mutant enzyme
-
-
Manually annotated by BRENDA team
strain NRC-1
-
-
Manually annotated by BRENDA team
gene cysS
-
-
Manually annotated by BRENDA team
prolyl-cysteinyl-tRNA synthetase, enzyme with dual specificity
-
-
Manually annotated by BRENDA team
gene cysS, canonical enzyme form CysRS
SwissProt
Manually annotated by BRENDA team
no activity in Archaeoglobus fulgidus
-
-
-
Manually annotated by BRENDA team
no activity in Methanococcus jannaschii
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
evolution
-
although the nucleotides in tRNA required for aminoacylation are conserved in evolution, bacterial aminoacyl-transfer RNA synthetases are unable to acylate eukaryote-specific tRNA. Whereas Escherichia coli CysRS cannot acylate human tRNACys, the fusion of a eukaryote-specific domain of human CysRS overcomes the cross-species barrier in human tRNACys. In addition to enabling recognition of the sequence differences in the tertiary core of tRNACys, the fused eukaryotic domain redirects the specificity of Escherichia coli CysRS from the A37 present in bacterial tRNACys to the G37 in mammals. The accuracy of codon recognition on the ribosome is also highly sensitive to the A37G transition in tRNACys
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
ATP + L-cysteine + tRNA1Cys
AMP + diphosphate + L-cysteinyl-tRNA1Cys
show the reaction diagram
-
substrate tRNA1Cys isoacceptor 1
-
-
?
ATP + L-cysteine + tRNA1CysA33U
AMP + diphosphate + L-cysteinyl-tRNA1CysA33U
show the reaction diagram
-
substrate tRNA1CysA33U isoacceptor 1, containing a A33U mutation
-
-
?
ATP + L-cysteine + tRNA2Cys
AMP + diphosphate + L-cysteinyl-tRNA2Cys
show the reaction diagram
-
substrate tRNA2Cys isoacceptor 2
-
-
?
ATP + L-cysteine + tRNA3Cys
AMP + diphosphate + L-cysteinyl-tRNA3Cys
show the reaction diagram
-
substrate tRNA3Cys isoacceptor 3
-
-
?
ATP + L-cysteine + tRNA3CysC20U/U21C/A44U/C46A/A47G
AMP + diphosphate + L-cysteinyl-tRNA3CysC20U/U21C/A44U/C46A/A47G
show the reaction diagram
-
substrate tRNA3CysC20U/U21C/A44U/C46A/A47G isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G mutation
-
-
?
ATP + L-cysteine + tRNA3CysC20U/U21C/A44U/C46A/A47G/G57A
AMP + diphosphate + L-cysteinyl-tRNA3CysC20U/U21C/A44U/C46A/A47G/G57A
show the reaction diagram
-
substrate tRNA3CysC20U/U21C/A44U/C46A/A47G/G57A isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G/G57A mutation
-
-
?
ATP + L-cysteine + tRNA3CysG57A
AMP + diphosphate + L-cysteinyl-tRNA3CysG57A
show the reaction diagram
-
substrate tRNA3CysG57A isoacceptor 3, containing a G57A mutation
-
-
?
ATP + L-cysteine + tRNA3CysU33A
AMP + diphosphate + L-cysteinyl-tRNA3CysU33A
show the reaction diagram
-
substrate tRNA3CysU33A isoacceptor 3, containing a U33A mutation
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
P21888
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
P21888
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-, Q9UWP2
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
tRNACys mutants: greatest loss of activity caused by mutation of a single nucleotide, occurs when the discriminator U73 is changed, mutations in the wobble nucleotide of the anticodon also cause reductions in the specificity constant of 3 orders of magnitude, while mutations in the other anticodon nucleotides causes lesser effects
-
-
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
P21888
enzyme is highly specific for L-cysteine and does not possess the editing activity characteristic for other tRNA synthetases
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
enzyme is highly specific for L-cysteine without performing an editing reaction, tightly bound zinc is the primary determinant of selectivity against non-cognate amino acids
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
two-step reaction mechanism, first step is the amino acid activation, formation of the intermediate Cys-AMP is dependent on tRNACys
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-, Q9UWP2
the canonical enzyme is not essential for viability of the archaeon Methanococcus maripaludis probably due to the dual-specific ProCys-tRNA synthetase
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
the attachment of cysteine to tRNACys by the class I cysteinyl-tRNA synthetase is flexible. The enzyme is capable of using either the 2' or 3'-hydroxyl group as the attachment site
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
a two-step process of amino acid activation as first step, and aminocylation as second step
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
the enzyme is rate-limited by release of aminoacyl-tRNA
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
Bacillus subtilis 168T
-
-
-
?
ATP + L-cysteine + tRNACys mutant
AMP + diphosphate + L-cysteinyl-tRNACys mutant
show the reaction diagram
-
tRNA substrate is a tRNACys with mutation at the core tertiary Levitt pair from wild-type G15.C48 to mutant G15.G48, leading to a higher activity
-
?
ATP + L-cysteine + tRNACysA36G
AMP + diphosphate + L-cysteinyl-tRNACysA36G
show the reaction diagram
-
relative activity compared to wild-type tRNACys as a substrate: 0.01
-
-
?
ATP + L-cysteine + tRNACysC35U
AMP + diphosphate + L-cysteinyl-tRNACysC35U
show the reaction diagram
-
relative activity compared to wild-type tRNACys as a substrate: 0.005
-
-
?
ATP + L-cysteine + tRNACysG15C/C48G
AMP + diphosphate + L-cysteinyl-tRNACysG15C/C48G
show the reaction diagram
-
relative activity compared to wild-type tRNACys as a substrate: 0.03
-
-
?
ATP + L-cysteine + tRNACysG34C
AMP + diphosphate + L-cysteinyl-tRNACysG34C
show the reaction diagram
-
relative activity compared to wild-type tRNACys as a substrate: 0.001
-
-
?
ATP + L-cysteine + tRNACysU73G
AMP + diphosphate + L-cysteinyl-tRNACysU73G
show the reaction diagram
-
relative activity compared to wild-type tRNACys as a substrate: 0.00002
-
-
?
ATP + L-cysteine + tRNAGln duoble-mutant
AMP + diphosphate + L-cysteinyl-tRNAGln double-mutant
show the reaction diagram
-
tRNA substrate is a tRNAGln with introduced tRNACys indentitiy nucleotides at the acceptor and anticodon ends and a core tertiary Levitt pair equivalent to tRNAGln of G15.G48, poor activity
-
?
ATP + L-cysteine + tRNAGln mutant
AMP + diphosphate + L-cysteinyl-tRNAGln mutant
show the reaction diagram
-
tRNA substrate is a tRNAGln with introduced tRNACys indentitiy nucleotides at the acceptor and anticodon ends and a core tertiary Levitt pair equivalent to tRNACys of G15.C48
-
?
ATP + L-proline + tRNAPro
AMP + diphosphate + L-prolyl-tRNAPro
show the reaction diagram
-
-, L-proline is the preferred substrate, two-step reaction mechanism, first step is the amino acid activation, intermediate Pro-AMP can be formed in absence of tRNAPro
-
?
L-Cysteinyl-tRNACys
Cysteine thiolactone + ?
show the reaction diagram
-
deacylation in which nucleophilic sulfur of the side chain of cysteine in Cys-tRNACys attacks its carboxyl carbon, synthesis of Cys-tRNACys and cyclization of cysteine to thiolactone occur in a single active site
-
-
additional information
?
-
-
ATP-diphosphate exchange
-
-
-
additional information
?
-
-
ATP-diphosphate exchange
-
-
-
additional information
?
-
-
ATP-diphosphate exchange
-
-
-
additional information
?
-
-
ATP-diphosphate exchange
-
-
-
additional information
?
-
-
ATP-diphosphate exchange
-
-
-
additional information
?
-
-
selenocysteine can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
selenocysteine can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
selenocysteine can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
selenocysteine can replace cysteine in ATP-diphosphate exchange, alanine shows minor activity in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
2-aminobutyric acid can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
2-aminobutyric acid can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
2-aminobutyric acid can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-
2-aminobutyric acid can replace cysteine in ATP-diphosphate exchange
-
-
-
additional information
?
-
-, Q9UWP2
the enzyme also performs the ATP-diphosphate exchange reaction
-
?
additional information
?
-
-
the enzyme performs the ATP-diphosphate exchange reaction with L-proline and L-cysteine, but with L-cysteine only in presence of tRNACys
-
?
additional information
?
-
-
probable regulation of enzyme activity by processing of polycistronic mRNA
-
?
additional information
?
-
-
Halobacterium sp. contains an unusual peptide that is unique to several halophile archaeal CysRS, which catalyze attachment of cysteine to tRNACys to generate the essential cysteinyl-tRNACys required for protein synthesis, deletion of the peptide reduces the catalytic efficiency of aminoacylation by a factor of 100 that largely results from a defect in kcat, rather than the Km for tRNACys, the acidity of the peptide is not important for the kcat of tRNA aminoacylation
-
-
-
additional information
?
-
-
in Methanosarcina mazei Cys-tRNACys is synthesized through a canonical cysteinyl-tRNA synthetase (CysRS) as well as via phosphoseryl-tRNACys synthesized by phosphoseryl-tRNA synthetase and this misacylated intermediate is then converted to Cys-tRNACys by Sep-tRNA:Cys-tRNA synthase via a pyridoxal phosphate-dependent mechanism. The genome of Methanosarcina mazei also features three distinct tRNACys isoacceptors, further indicating the unusual and complex nature of Cys-tRNACys synthesis in this organism
-
-
-
additional information
?
-
Bacillus subtilis 168T
-
probable regulation of enzyme activity by processing of polycistronic mRNA
-
?
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
P21888
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-
-
-
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
-, Q9UWP2
the canonical enzyme is not essential for viability of the archaeon Methanococcus maripaludis probably due to the dual-specific ProCys-tRNA synthetase
-
?
ATP + L-cysteine + tRNACys
AMP + diphosphate + L-cysteinyl-tRNACys
show the reaction diagram
Bacillus subtilis 168T
-
-
-
?
ATP + L-proline + tRNAPro
AMP + diphosphate + L-prolyl-tRNAPro
show the reaction diagram
-
-
-
?
additional information
?
-
Bacillus subtilis, Bacillus subtilis 168T
-
probable regulation of enzyme activity by processing of polycistronic mRNA
-
?
COFACTOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
ATP
-
dependent on, optimal concentration is 3 mM
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Ca2+
-
15% of the efficiency of Mg2+ in activation of ATP-diphosphate exchange
Co2+
-
13% of the efficiency of Mg2+ in activation of ATP-diphosphate exchange
Co2+
-
can substitute for Zn2+
KCl
-
required, optimal cocentration is 150 mM
Mg2+
-
optimal Mg2+/ATP ratio is 2.5; required for ATP-diphosphate exchange
Mg2+
-
inhibition above 15 mM; Km: 3.6 mM; optimal concentration: 10 mM; required for ATP-diphosphate exchange
Mg2+
-
inhibited by free Mg2+; optimal Mg2+/ATP ratio is 2.5; required for ATP-diphosphate exchange
Mg2+
-
dependent on, optimal concentration is 5 mM
Mg2+
-
-
Mn2+
-
9% of the efficiency of Mg2+ in activation of ATP-diphosphate exchange
Mn2+
-
can partially replace Mg2+ in activation of cysteine, optimal concentration is 5 mM
Zn2+
-
6% of the efficiency of Mg2+ in activation of ATP-diphosphate exchange
Zn2+
P21888
1 ion bound at the base of the active site cleft, important for tRNA substrate specificity, single direct interaction with cysteine thiolate substrate
Zn2+
-
can be substituted by Co2+, tightly bound, 1 ion per enzyme molecule, bound at the base of the active site cleft, primary determinant of selectivity against non-cognate amino acids
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
2'-Deoxy-ATP
-
-
2-L-aminobutyric acid
-
-
5'-O-[N-(L-cysteinyl)sulfamoyl] adenosine
-
i.e. Cys-AMS, a cysteinyl adenylate analogue
ADP
-
noncompetitive
alpha,beta-CH2-ATP
-
competitive
AMP
-
noncompetitive
ATP
-
free ATP inhibits
ATP
-
at high concentrations
ATP
-
free ATP inhibits
Ba2+
-
10 mM, 43% inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
Ca2+
-
10 mM, 28% inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
Co2+
-
10 mM, complete inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
Cu2+
-
10 mM, complete inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
Cys-AMP
-
inhibits deacylation of Cys-tRNACys
-
cysteamine
-
competitive
cysteine
-
inhibits deacylation of Cys-tRNACys
cysteine
-
; at high concentrations
D-Cysteine
-
weak, non-competitive
diphosphate
-
at high concentrations
glutathione
-
reduced
GTP
-
noncompetitive
Hg2+
-
10 mM, complete inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
iodoacetamide
-
partially
L-cysteic acid
-
-
L-cysteine
-
competitive inhibition of prolylation, a 40fold excess over L-proline concentration reduces the prolylation activity by 80%, no inhibition of mutant P100A
L-homocysteine
-
-
L-homocysteine
-
-
L-proline
-
competitive inhibition of cysteinylation, a 40fold excess over L-cysteine concentration reduces the cysteinylation activity by over 80%, no inhibition of mutant E103A
L-selenocysteine
-
-
O-acetylserine
-
-
p-chloromercuribenzoate
-
-
p-chloromercuribenzoate
-
-
phenylhydrazine
-
weak
S-methyl-L-cysteine
-
competitive
Sodium tripolyphosphate
-
-
Sulfhydryl group reagents
-
-
-
Zn2+
-
10 mM, complete inhibition in presence of 10 mM Mg2+, no activation in absence of Mg2+
Mg2+
-
free Mg2+ inhibits
additional information
-
activity is not affected by addition of diphosphate
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
methionine
-
stimulates
tRNACys
-
promotes its aminoacylation with L-cysteine
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
10
-
2-aminobutyric acid
-
ATP-diphosphate exchange
10
-
2-aminobutyric acid
-
ATP-diphosphate exchange
0.049
-
ATP
-
aminoacylation reaction with L-proline, recombinant wild-type enzyme, pH 7.0, 70C
0.057
-
ATP
-
ATP-diphosphate exchange, pH 5.12, 37C
0.06
-
ATP
-
aminoacylation reaction with L-cysteine, recombinant wild-type enzyme, pH 7.0, 70C
0.083
-
ATP
-
ATP-diphosphate exchange, 17C, pH 7.5
0.089
-
ATP
-
ATP-diphosphate exchange, 19C, pH 7.5
0.094
-
ATP
-
ATP-diphosphate exchange, 21C, pH 7.5
0.095
-
ATP
-
ATP-diphosphate exchange, pH 5.6, 37C
0.1
-
ATP
-
ATP-diphosphate exchange, 23C, pH 7.5
0.111
-
ATP
-
ATP-diphosphate exchange, 25C, pH 7.5
0.147
-
ATP
-
ATP-diphosphate exchange, pH 6.04, 37C
0.172
-
ATP
-
ATP-diphosphate exchange, pH 6.53, 37C
0.22
-
ATP
-
ATP-diphosphate exchange reaction, recombinant His-tagged Co2+-wild-type enzyme
0.25
-
ATP
-
ATP-diphosphate exchange reaction, recombinant His-tagged Zn2+-wild-type enzyme
0.25
-
ATP
-
pH 8.0, wild-type enzyme, ATP-diphosphate exchange
0.29
-
ATP
-
ATP-diphosphate exchange reaction, recombinant Zn2+-wild-type enzyme
0.31
-
ATP
-
pH 8.0, mutant enzyme DELTA288-461, ATP-diphosphate exchange
0.338
-
ATP
-
30C, pH 7.0, wild-type enzyme
0.39
-
ATP
-
ATP-diphosphate exchange reaction with L-cysteine, recombinant wild-type enzyme, pH 7.2, 70C
0.412
-
ATP
-
30C, pH 7.0, mutant enzyme V27E
0.465
-
ATP
-
ATP-diphosphate exchange reaction with L-proline, recombinant wild-type enzyme, pH 7.2, 70C
0.77
-
ATP
-
ATP-diphosphate exchange reaction, recombinant W205Y mutant enzyme
0.8
-
ATP
-
pH 7.5, 25C
1.15
-
ATP
-
ATP-diphosphate exchanhge
1.18
-
ATP
-
pH 8.0, mutant enzyme DELTA328-461, ATP-diphosphate exchange
1.33
-
ATP
-
ATP-diphosphate exchange
0.03
-
Cys
-
-
0.045
-
Cys
-
ATP-diphosphate exchange
0.06
-
Cys
-
ATP-diphosphate exchange
0.062
-
Cys
-
ATP-diphosphate exchange
0.17
-
Cys
-
ATP-diphosphate exchange
0.16
-
diphosphate
-
-
0.3
-
diphosphate
-
ATP-diphosphate exchange
1
-
diphosphate
-
ATP-diphosphate exchange
0.0125
-
L-Cys
-
ATP-diphosphate exchange
0.0072
-
L-cysteine
-
30C, pH 7.0, wild-type enzyme
0.0097
-
L-cysteine
-, Q9UWP2
recombinant wild-type enzyme, pH 7.0, 37C
0.0103
-
L-cysteine
-
recombinant His-tagged deletion mutant, amino acid activation step
0.011
-
L-cysteine
-
pH 7.5, 25C
0.0203
-
L-cysteine
-
recombinant His-tagged wild-type full-length enzyme, amino acid activation step
0.022
-
L-cysteine
-
aminoacylation reaction, recombinant wild-type enzyme, pH 7.0, 70C
0.022
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant Zn2+-wild-type enzyme
0.0282
-
L-cysteine
-
30C, pH 7.0, mutant enzyme V27E
0.029
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant His-tagged Co2+-wild-type enzyme
0.031
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant His-tagged Zn2+-wild-type enzyme
0.09
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant wild-type enzyme, pH 7.2, 70C
3.61
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant W205Y mutant enzyme
0.027
-
L-proline
-
aminoacylation reaction, recombinant wild-type enzyme, pH 7.0, 70C
0.285
-
L-proline
-
ATP-diphosphate exchange reaction, recombinant wild-type enzyme, pH 7.2, 70C
0.05
-
selenocysteine
-
ATP-diphosphate exchange
0.05
-
selenocysteine
-
ATP-diphosphate exchange
0.063
-
selenocysteine
-
ATP-diphosphate exchange
0.08
-
tRNA1Cys
-
value above, substrate tRNA1Cys isoacceptor 1
-
0.09
-
tRNA1CysA33U
-
value above, substrate tRNA1Cys isoacceptor 1, containing a A33U mutation
-
0.16
-
tRNA2Cys
-
value above, substrate tRNA2Cys isoacceptor 2
-
0.14
-
tRNA3Cys
-
value above, substrate tRNA3Cys isoacceptor 3
-
0.11
-
tRNA3CysC20U/U21C/A44U/C46A/A47G
-
value above, substrate tRNA3Cys isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G mutation
-
0.09
-
tRNA3CysC20U/U21C/A44U/C46A/A47G/G57A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G/G57A mutation
-
0.12
-
tRNA3CysG57A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a G57A mutation
-
0.13
-
tRNA3CysU33A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a U33A mutation
-
0.00035
-
tRNACys
-
aminoacylation reaction, recombinant Zn2+-wild-type enzyme
0.0004
-
tRNACys
-
aminoacylation reaction, recombinant His-tagged Co2+-wild-type enzyme
0.0005
-
tRNACys
-
mutant enzyme E354A, wild-type tRNACys(G15-G48)
0.00058
-
tRNACys
-
mutant enzyme E354Q, wild-type tRNACys(G15-G48)
0.00064
-
tRNACys
-
30C, pH 7.0, wild-type enzyme
0.0009
-
tRNACys
-
recombinant His-tagged wild-type full-length enzyme, amino acid activation step
0.00092
-
tRNACys
-
30C, pH 7.0, mutant enzyme V27E
0.00116
-
tRNACys
-
37C, wild-type enzyme
0.00116
-
tRNACys
-
wild-type enzyme, wild-type tRNACys(G15-G48)
0.0012
-
tRNACys
-
aminoacylation reaction, recombinant His-tagged Zn2+-wild-type enzyme
0.0012
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D417A/E420A
0.0013
-
tRNACys
-
pH 7.5, 40C, recombinant wild-type enzyme
0.0015
-
tRNACys
-
pH 7.0, 37C
0.0017
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D239A/D240A
0.0023
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D435A/D436A
0.0045
-
tRNACys
-
recombinant His-tagged deletion mutant, amino acid activation step
0.0066
-
tRNACys
-
aminoacylation reaction, recombinant W205Y mutant enzyme
0.0088
-
tRNACys
-
mutant enzyme N351A, wild-type tRNACys(G15-G48)
0.0177
-
tRNACys
-
mutant enzyme N351D, mutant tRNACys(G15-C48)
0.0179
-
tRNACys
-
mutant enzyme E354Q, wild-type tRNACys(G15-G48)
0.0185
-
tRNACys
-
mutant enzyme N351D, wild-type tRNACys(G15-G48)
0.0332
-
tRNACys
-
mutant enzyme N351A, mutant tRNACys(G15-C48)
0.0358
-
tRNACys
-
wild-type enzyme, mutant tRNACys(G15-C48)
0.0435
-
tRNACys
-
mutant enzyme E354A, mutant tRNACys(G15-C48)
0.08
-
tRNACys
-
pH 7.5, 25C
0.095
-
tRNACys
-
aminoacylation reaction, recombinant C28S/C209S mutant enzyme
0.00159
-
L-selenocysteine
-
ATP-diphosphate exchange
additional information
-
additional information
-
Km-values of mutant tRNA molecules
-
additional information
-
additional information
-
dependence of Km value on pH and temperature
-
additional information
-
additional information
-
Km for ATP and tRNA is similar in normal and cystinotic cells
-
additional information
-
additional information
-
tRNAGln mutant substrates
-
additional information
-
additional information
-
thermodynamics, single turnover and burst kinetics of CysRS, steady-state and transient kinetic analyses of class I CysRS, the enzyme is rate-limited by release of aminoacyl-tRNA, recombinant His-tagged enzyme, overview
-
additional information
-
additional information
-
steady-state, single turnover, and multiple turnover kinetics of recombinant His-tagged wild-type full-length enzyme, and recombinant His-tagged wild-type deletion mutant, kinetic analysis of the two reaction steps, overview
-
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.3
-
ATP
-
ATP-diphosphate exchange reaction, recombinant W205Y mutant enzyme
0.34
-
ATP
-
pH 8.0, mutant enzyme DELTA328-461, ATP-diphosphate exchange
0.48
-
ATP
-
30C, pH 7.0, mutant enzyme V27E
2
-
ATP
-
pH 8.0, mutant enzyme DELTA288-461, ATP-diphosphate exchange
4.4
-
ATP
-
30C, pH 7.0, wild-type enzyme
22.6
-
ATP
-
pH 8.0, wild-type enzyme, ATP-diphosphate exchange
57
-
ATP
-
ATP-diphosphate exchange reaction, recombinant His-tagged Co2+-wild-type enzyme
91
-
ATP
-
ATP-diphosphate exchange reaction, recombinant Zn2+-wild-type enzyme
142
-
ATP
-
ATP-diphosphate exchange reaction, recombinant His-tagged Zn2+-wild-type enzyme
0.02
-
L-cysteine
-
aminoacylation reaction, recombinant wild-type enzyme, pH 7.0, 70C
0.04
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant wild-type enzyme, pH 7.2, 70C
0.28
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant W205Y mutant enzyme
0.86
-
L-cysteine
-
30C, pH 7.0, mutant enzyme V27E
2.2
-
L-cysteine
-, Q9UWP2
recombinant wild-type enzyme, pH 7.0, 37C
2.5
-
L-cysteine
-
steady-state
4.8
-
L-cysteine
-
30C, pH 7.0, wild-type enzyme
10.3
-
L-cysteine
-
recombinant His-tagged wild-type full-length enzyme, amino acid activation step
18.1
-
L-cysteine
-
recombinant His-tagged deletion mutant, amino acid activation step
35
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant His-tagged Co2+-wild-type enzyme
79
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant Zn2+-wild-type enzyme
100
-
L-cysteine
-
ATP-diphosphate exchange reaction, recombinant His-tagged Zn2+-wild-type enzyme
0.09
-
L-proline
-
aminoacylation reaction, recombinant wild-type enzyme, pH 7.0, 70C
0.017
-
thiolactone formation
-
-
-
0.17
-
tRNA1Cys
-
value above, substrate tRNA1Cys isoacceptor 1
-
0.16
-
tRNA1CysA33U
-
value above, substrate tRNA1Cys isoacceptor 1, containing a A33U mutation
-
0.06
-
tRNA2Cys
-
value above, substrate tRNA2Cys isoacceptor 2
-
0.03
-
tRNA3Cys
-
value above, substrate tRNA3Cys isoacceptor 3
-
0.14
-
tRNA3CysC20U/U21C/A44U/C46A/A47G
-
value above, substrate tRNA3Cys isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G mutation
-
0.18
-
tRNA3CysC20U/U21C/A44U/C46A/A47G/G57A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a C20U/U21C/A44U/C46A/A47G/G57A mutation
-
0.09
-
tRNA3CysG57A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a G57A mutation
-
0.05
-
tRNA3CysU33A
-
value above, substrate tRNA3Cys isoacceptor 3, containing a U33A mutation
-
0.005
-
tRNACys
-
aminoacylation reaction, recombinant C28S/C209S mutant enzyme
0.007
-
tRNACys
-
aminoacylation reaction, recombinant W205Y mutant enzyme
0.014
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D417A/E420A
0.03
-
tRNACys
-
mutant enzyme N351D, mutant tRNACys(G15-C48)
0.03
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D435A/D436A
0.036
-
tRNACys
-
mutant enzyme E354A, mutant tRNACys(G15-C48)
0.06
-
tRNACys
-
pH 7.5, 40C, recombinant mutant D239A/D240A
0.074
-
tRNACys
-
mutant enzyme E354Q, mutant tRNACys(G15-C48)
0.094
-
tRNACys
-
mutant enzyme N351D, wild-type tRNACys(G15-G48)
0.1
-
tRNACys
-
mutant enzyme E354A, wild-type tRNACys(G15-G48)
0.11
-
tRNACys
-
pH 7.5, 40C, recombinant wild-type enzyme
0.14
-
tRNACys
-
mutant enzyme E354Q, wild-type tRNACys(G15-G48)
0.2
-
tRNACys
-
recombinant His-tagged deletion mutant, amino acid activation step
0.39
-
tRNACys
-
mutant enzyme N351A, mutant tRNACys(G15-C48)
0.45
-
tRNACys
-
wild-type enzyme, mutant tRNACys(G15-C48)
0.5
-
tRNACys
-
aminoacylation reaction, recombinant His-tagged Co2+-wild-type enzyme
0.59
-
tRNACys
-
mutant enzyme N351A, wild-type tRNACys(G15-G48)
0.68
-
tRNACys
-
30C, pH 7.0, mutant enzyme V27E
0.9
-
tRNACys
-
aminoacylation reaction, recombinant Zn2+-wild-type enzyme
2.3
-
tRNACys
-
recombinant His-tagged wild-type full-length enzyme, amino acid activation step
2.46
-
tRNACys
-
37C, wild-type enzyme
2.46
-
tRNACys
-
wild-type enzyme, wild-type tRNACys(G15-G48)
2.5
-
tRNACys
-
aminoacylation reaction, recombinant His-tagged Zn2+-wild-type enzyme
2.9
-
tRNACys
-
30C, pH 7.0, wild-type enzyme
2.9
-
tRNACys
-
steady-state
3
6
tRNACys
-
pH 7.0, 37C
3.47
-
tRNACys
-
pH 7.0, 37C
4.8
-
L-proline
-
ATP-diphosphate exchange reaction, recombinant wild-type enzyme, pH 7.2, 70C
additional information
-
additional information
-
turnover numbers of mutant tRNA molecules
-
additional information
-
additional information
-
tRNAGln mutant substrates
-
SPECIFIC ACTIVITY [µmol/min/mg]
SPECIFIC ACTIVITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
0.22
-
-
purified enzyme
1.2
-
-
enzyme form CRS-1
3.29
-
-
enzyme form CRS-2
6.25
-
-
enzyme form CRS-3
additional information
-
-
-
additional information
-
-
-
additional information
-
-
-
additional information
-
-
-
additional information
-
-
-
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
7
-
-
aminoacylation assay at
7
-
-
assay at
7
-
-, Q9UWP2
assay at
7.2
-
-
ATP-diphosphate exchange assay at
7.4
-
-
-
7.5
-
-
assay at
7.5
-
-
assay at
7.5
-
-
assay at
8
-
-
ATP-diphosphate exchange
pH RANGE
pH RANGE MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
7
9
-
pH 7.0: about 30% of maximal activity, pH 9.0: about 50% of maximal activity, ATP-diphosphate exchange
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
25
-
-
assay at
37
-
-
assay at
37
-
-, Q9UWP2
assay at
37
-
-
assay at
40
-
-
assay at
SOURCE TISSUE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
SOURCE
-
cultured; from patients with cystinosis
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
Borrelia burgdorferi (strain ATCC 35210 / B31 / CIP 102532 / DSM 4680)
Coxiella burnetii (strain RSA 493 / Nine Mile phase I)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
42320
-
-
predicted from cDNA
52000
-
-
-
54000
-
-
gel filtration
61000
-
-
gel filtration
82000
-
-
gel filtration, mitochondrial enzyme
122000
-
-
gel filtration, enzyme form, CRS-1
127000
-
-
sucrose density gradient centrifugation, enzyme form CRS-2
230000
240000
-
gel filtration, PAGE on gels of various porosities, enzyme form CRS-2
262000
-
-
gel filtration, cytoplasmic enzyme
270000
300000
-
gel filtration, enzyme form CRS-3
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 52280, deduced from nucleotide sequence
?
-
x * 92000, enzyme form I, SDS-PAGE, x * 86000, enzyme form II, SDS-PAGE
dimer
-
2 * 30000, SDS-PAGE + 8 M urea
dimer
-
2 * 120000, SDS-PAGE in presence of 2-mercaptoethanol, enzyme form CRS-2
dimer
-
2 * 87000, recombinant His-tagged wild-type full-length enzyme, SDS-PAGE, 2 * 76000, recombinant His-tagged deletion mutant enzymes, SDS-PAGE
dimer
-
gel filtration, 2 * 42324
monomer
-
1 * 54000, SDS-PAGE
monomer
-
1 * 52000
monomer
P21888
-
additional information
-
the enzyme forms a class I ternary complex with EF-Tu, overview
additional information
-
human CysRS does not associate with the multi-synthetase complex, yet contains an eukaryotic-specific C-terminal extension that follows the tRNA anticodon-binding domain, the C-terminal extension is not involved in dimer formation, but is important for stable binding of tRNACys in the aminoacylation reaction step
Crystallization/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
hanging drop vapor diffusion method, crystals of enzyme bound to tRNACys at a resolution of 2.3 A
-
purified enzyme complexed with ATP and cysteine, hanging-drop vapour diffusion from ammonium sulfate precipitant containing solution, X-ray diffraction structure analysis at 2.7 A resolution
-
purified recombinant free enzyme or cysteine and ATP complexed enzyme, hanging-drop vapour diffusion method, 17C, 0.001 ml protein solution: 6 mg/ml enzyme, 10 mM HEPES, pH 7.4, 50 mM NaCl, 1 mM DTT, 5 mM MgCl2, 5 mM ATP, 10 mM cysteine, plus equal volume of reservoir solution: 0.1 M sodium cacodylate, pH 6.5, 15-17% PEG 8000, 0.2 M magnesium acetate, 2% tert-butanol, 1-3 weeks, cysteine but not ATP is required for crystal growth, X-ray diffraction structure determination at 2.3-3.0 A resolution, and analysis
P21888
pH STABILITY
pH STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
6
9
-
37C, in presence of 10% glycerol, stable for at least 15 min
TEMPERATURE STABILITY
TEMPERATURE STABILITY MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
37
-
-
pH 6-9, in presence of 10% glycerol, stable for at least 15 min
GENERAL STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
instability of the enzyme without the halophilic-specific peptide
-
STORAGE STABILITY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
-20C, 50% glycerol
-
-20C, 50% glycerol, 1 mM DTE, at least 6 months, no loss of activity
-
4C, 50% glycerol, 0.5 mM DTT, 3 mM L-cysteine
-
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
partial
-
recombinant from overexpressing strain JM109
-
recombinant His-tagged wild-type enzyme and halophilic-specific peptide mutant enzyme variants from Escherichia coli strain BL21(DE3)
-
recombinant C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli to homogeneity
-
recombinant His-tagged wild-type and mutants from overexpressing Escherichia coli BL21
-
using Ni-NTA chromatographay
-
8300fold from liver, 2 different forms, to homogeneity
-
multiple molecular forms: CRS-1, CRS-2, CRS-3
-
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
the gene encoding the enzyme is expressed as a polycistronic gltX-cysE-cysS transcript, which is processed to a cysE-cysS transcript, regulatory implications
-
overexpression in strain JM109
-
expression of His-tagged wild-type enzyme and halophilic-specific peptide mutant enzyme variants in Escherichia coli strain BL21(DE3)
-
gene cysS, stable expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli
-
gene proS, expression of wild-type and mutant enzymes in Escherichia coli BL21
-
overexpression of the wild-type enzyme in Escherichia coli BL21(DE3) as N-terminally HIs-tagged protein
-, Q9UWP2
expressed as a His-tagged fusion protein in Escherichia coli
-
ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
C209S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.0031% of the wild-type ratio
C28S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.012% of the wild-type ratio
C28S/C209S
-
site-directed mutagenesis, no ATP-diphosphate exchange activity, residual aminoacylation activity
C28S/C209S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.0026% of the wild-type ratio
C28S/C209S/H234N/E238Q
-
site-directed mutagensis, mutation of all zinc binding ligands, complete loss of bound zinc, weak ability to bind serine, thus loss of amino acid substrate discrimination ability
C28S/C209S/H234N/E238Q
-
aminoacylation of tRNACys is not detectable
C36S/C214S/C244S
-
site-directed mutagenesis, activity is similar to the wild-type enzyme, but the affinity for cysteine binding is increased
DELTA288-461
-
the ratio of turnover number to Km-value for ATP in ATP-diphosphate exchange is 7% of the wild-type ratio
DELTA328-461
-
the ratio of turnover number to Km-value for ATP in ATP-diphosphate exchange is 0.32% of the wild-type ratio, aminoacylation of tRNACys is not detectable
E354Q
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 10% of the wild-type ratio
H206S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 60% of the wild-type ratio
H224N/H235N
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.042% of the wild-type ratio
H224S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 5.7% of the wild-type ratio
H234N/E238Q
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.0059% of the wild-type ratio
H234N/E238Q/H224N/H235N
-
aminoacylation of tRNACys is not detectable
H234S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.015% of the wild-type ratio
H235S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.38% of the wild-type ratio
H238S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 30% of the wild-type ratio
N351D
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.25% of the wild-type ratio
V27E
-
mutation does not affect the discrimination of the enzyme for serine. 4fold increase in Km-value for cysteine and 9fold reduction of turnover number for ATP
W205F
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 0.55% of the wild-type ratio
W205Y
-
site-directed mutagenesis, highly reduced activity, highly increased Km for cysteine
D239A/D240A
-
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
D417A/E420A
-
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
E103A
-
site-directed mutagenesis, unaltered thermostability, no remaining prolylation activity, 5% remaining cysteinylation activity compared to the wild-type enzyme
H256S
-
the ratio of turnover number to Km-value of aminoacylation of tRNACys is 10% of the wild-type ratio
additional information
-
construction of a Co2+-substituted wild-type enzyme, similar properties as the Zn2+-wild-type enzyme but slightly reduced activity
additional information
-
construction of a fusion of a eukaryote-specific domain of human CysRS enabling recognition of the sequence differences in the tertiary core of tRNACys. The fused eukaryotic domain redirects the specificity of Escherichia coli CysRS from the A37 present in bacterial tRNACys to the G37 in mammals
D435A/D436A
-
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
additional information
-
deletion of the halophilic-specific peptide reduces the catalytic efficiency of aminoacylation by a factor of 100 that largely results from a defect in kcat, rather than the Km for tRNACys, maintaining the peptide length but substituting acidic residues in the peptide with neutral or basic residues has no major deleterious effect, suggesting that the acidity of the peptide is not important for the kcat of tRNA aminoacylation, construction of point mutants and deletion mutants, e.g. deletion mutant DELTA193-212, overview
additional information
-
construction of a DELTAC mutant of the enzyme that terminates at V642, removing residues 643 to 748 from the full-length enzyme, the mutant enzymes shows increased activity in the first amino acid activation step and reduced activity in the second aminoacylation step compared to the wild-type full-length enzyme
P100A
-
site-directed mutagenesis, unaltered thermostability, loss of 90% cysteinylation activity, unaltered prolylation activity compared to the wild-type enzyme
additional information
-, Q9UWP2
construction of a cysS gene disruption knockout mutant, which is viable under normal growth conditions
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
medicine
-
in order to evaluate genetic associations between candidate genes involved in oxidative stress and multiple system atrophy (MSA) 119 Japanese patients with MSA and 123 controls are examined and single-nucleotide polymorphisms are genotyped. Results revealed genetic associations of cysteinyl-tRNA synthetase, solute carrier family 1A4, sequestosome 1, and eukaryotic translation initiation factor 4E-binding protein 1 with MSA
molecular biology
-
Mycoplasmas hyopneumoniae are collected from bronchial alveolar lavage samples of infected pigs 28 days postinfection and compared to broth-grown cells using microarrays. During lung infection, the analysis indicates that 79 genes are differentially expressed. Of the down-regulated genes, 28 of 46 (61%) lacked an assigned function, in comparison to 21 of 33 (63%) of up-regulated genes. Cysteinyl-tRNA synthetase is down-regulated in vivo