Information on EC 4.4.1.15 - D-cysteine desulfhydrase

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

EC NUMBER
COMMENTARY hide
4.4.1.15
-
RECOMMENDED NAME
GeneOntology No.
D-cysteine desulfhydrase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
D-cysteine + H2O = sulfide + NH3 + pyruvate
show the reaction diagram
-
-
-
-
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C-S-bond cleavage
elimination of H2S or RSH
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Cysteine and methionine metabolism
-
-
cysteine metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
D-cysteine sulfide-lyase (deaminating; pyruvate-forming)
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CAS REGISTRY NUMBER
COMMENTARY hide
84012-74-8
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
green alga, strain 211-8b, formerly Chlorella pyrenoidosa strain 211-8b
-
-
Manually annotated by BRENDA team
-
UniProt
Manually annotated by BRENDA team
strain W3110 deltatrpED102/F'deltatrpED102
-
-
Manually annotated by BRENDA team
tobacco, var. Samsun
-
-
Manually annotated by BRENDA team
strain CU 1462/7
-
-
Manually annotated by BRENDA team
strain CU 1462/7
-
-
Manually annotated by BRENDA team
strain CR 1-1
-
-
Manually annotated by BRENDA team
strain UW4
-
-
Manually annotated by BRENDA team
bony best variety
UniProt
Manually annotated by BRENDA team
strain PCC7942
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
the enzyme structurally belongs to the fold type II pyridoxal 5'-phosphate-dependent enzyme family
metabolism
physiological function
additional information
active site structure analysis and comparisons, residue Tr287 is important for catalysis, while His80 and Tyr261 may not be directly involved in the degradation of D-Cys, overview
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3-chloro-D-alanine + H2O
pyruvate + NH3 + Cl-
show the reaction diagram
3-chloro-D-alanine + thioglycolic acid
S-carboxymethyl-D-cysteine
show the reaction diagram
D-allocystathionine + H2O
?
show the reaction diagram
-
-
-
-
?
D-cysteine + H2O
sulfide + NH3 + pyruvate
show the reaction diagram
D-cystine + H2O
?
show the reaction diagram
-
-
-
-
?
DL-lanthionine + H2O
?
show the reaction diagram
-
-
-
-
?
DL-selenocysteine + H2O
selenide + L-alanine
show the reaction diagram
-
very poor substrate
-
-
?
DL-selenocystine + H2O
?
show the reaction diagram
-
-
-
-
?
L-cysteine + H2O
sulfide + NH3 + pyruvate
show the reaction diagram
L-selenocystine + H2O
selenide + L-alanine
show the reaction diagram
-
-
-
-
?
O-acetyl-D-serine + H2O
?
show the reaction diagram
O-acetyl-D-serine + sulfide
D-cysteine + ?
show the reaction diagram
O-methyl-DL-serine + H2O
?
show the reaction diagram
-
poor substrate
-
-
?
S-benzyl-D-cysteine + H2O
benzyl hydrosulfide + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-carboxymethyl-D-cysteine + H2O
mercaptoacetic acid + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-ethyl-D-cysteine + H2O
ethanethiol + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-methyl-D-cysteine + H2O
methanethiol + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-n-butyl-D-cysteine + H2O
butane-1-thiol + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-n-propyl-D-cysteine + H2O
propane-1-thiol + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
S-phenyl-D-cysteine + H2O
benzenthiol + pyruvate + NH3
show the reaction diagram
-
-
-
-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
D-cysteine + H2O
sulfide + NH3 + pyruvate
show the reaction diagram
additional information
?
-
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
pyridoxal 5'-phosphate
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
(1-aminoethyl)phosphonic acid
-
-
aminooxy acetic acid
-
at 10 microM, IC50 value is 0.0305 mM
aminooxyacetic acid
D-penicillamine
-
-
hydroxylamine
L-cysteine
-
mixed type inhibition
L-Penicillamine
-
-
p-chloromercuribenzoate
-
-
phenylhydrazine
-
-
pyruvate
additional information
-
no inhibition with EDTA and substrates
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
dithiothreitol
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.9 - 0.91
3-chloro-D-alanine
0.29
D-allocystathionine
-
30C, pH 8.0
0.05 - 0.34
D-Cysteine
0.27
D-Cystine
-
30C, pH 8.0
0.11
DL-lanthionine
-
30C, pH 8.0, based on the concentration of the D-isomer, Escherichia coli
0.04
DL-selenocystine
-
30C, pH 8.0, based on the concentration of the D-isomer, Escherichia coli
0.28
S-benzyl-D-cysteine
-
30C, pH 8.0
0.83
S-carboxymethyl-D-cysteine
-
30C, pH 8.0
0.42
S-ethyl-D-cysteine
-
30C, pH 8.0
0.5
S-methyl-D-cysteine
-
30C, pH 8.0
0.24
S-n-butyl-D-cysteine
-
30C, pH 8.0
0.23
S-n-propyl-D-cysteine
-
30C, pH 8.0
0.22
S-phenyl-D-cysteine
-
30C, pH 8.0
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
281
3-chloro-D-alanine
Escherichia coli
-
37C, pH 8.0
6 - 13800
D-Cysteine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0033
aminooxyacetic acid
value bases on non-linear regression
0.53
L-cysteine
-
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0305
aminooxy acetic acid
Arabidopsis thaliana
-
at 10 microM, IC50 value is 0.0305 mM
0.0159
hydroxylamine
Arabidopsis thaliana
-
at 10 microM, IC50 value is 0.0159 mM
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.001594
-
D-cysteine desulfhydrase activity in WT
0.012
-
growth of the bacteria with methionine as sulfur source
0.0123
-
after partial purification
0.018
-
growth of the bacteria with L-cysteine as sulfur; growth of the bacteria with Na2S2O3 and p-nitrophenlysulfate as sulfur; growth of the bacteria with Na2S2O3 as sulfur source
0.02
-
growth of the bacteria with MgSO4 and p-nitrophenlysulfate as sulfur source
0.022
-
growth of the bacteria with MgSO4 as sulfur source
0.03
-
growth of the bacteria with MgSO4 and Na2S2O3 as sulfur source
0.035
-
growth of the bacteria with reduced glutathione as sulfur source
0.0356
-
D-cysteine desulfhydrase activity in mutant E295S
0.1475
-
D-cysteine desulfhydrase activity in double mutant E295S/L322T
0.21
-
purified enzyme, 37C, pH 9.0
0.6653
D-cysteine desulfhydrase activity in mutant T386L. Changing the threonine 386 residue alone reduces the activity of the enzyme substantially of about 11.5% of the native recombinant enzyme, although it does not cause complete loss of activity.
5.784
D-cysteine desulfhydrase activity in wild-type
13
-
purified enzyme, 30C, pH 8.0, with D-cysteine as substrate
56.3
-
purified enzyme, 30C, pH 8.0, with 3-chloro-D-alanine as substrate
additional information
activity below detection, D-cysteine desulfhydrase activity in double mutant S358E/T386L; activity below detection, D-cysteine desulfhydrase activity in mutant S358E. Serine at 358 residue is essential for D-cysteine desulfhydrase activity.
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.6
with estimated 6.7 (pKa1) and 8.9 (pKa2)
8.5 - 9
9
-
alpha,beta-elimination
9.5
-
beta-replacement
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
37
-
assay at
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
-
loss of activity at 50C, no activity at 60C. The enzyme is also very sensitive to freezing. One freeze-thaw cycle leads to a loss of activity of 75%.
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.34
-
mature protein without N-terminal extension, calculated from amino acid sequence
7.2
-
calculated from amino acid sequence
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
collected at the breaker stage, which is defined as the stage where first spot of pink/red color appears at the blossom end
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35000
-
SDS-PAGE, gel filtration
35150
-
calculated from amino acid sequence including N-terminal methionine
36500
-
SDS-PAGE
41700
-
monomer, mature protein without N-terminal extension, calculated from amino acid sequence
43900
-
monomer, calculated from amino acid sequence
51480
recombinant protein is determined by mass spectrometry
70000
-
gel filtration
74000
-
HPLC gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
-
x * 42000, SDS-PAGE, unprocessed protein; x * 43000, SDS-PAGE, mature protein; x * 44000, SDS-PAGE, post-translationally modified protein
additional information
the polypeptide fold of the enzyme consists of a small domain (residues 48161) and a large domain (residues 1-47 and 162-328)
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified enzyme in complex with D-Cys, beta-chloro-D-alanine, 1-amino-1-carboxy cyclopropane, D-Ser, L-Ser, D-cycloserine, and L-cycloserine, X-ray diffraction structure determination and analysis at 1.7-2.6 A resolution, modeling
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50
The enzyme is found to be very stable with almost 80% of its activity still remaining at 50C
60
-
no loss of activity within 30 min incubation, 0.1 M phosphate buffer, pH 8.0
65
-
11% loss of activity within 30 min incubation, 0.1 M phosphate buffer, pH 8.0
70
-
24% loss of activity within 30 min incubation, 0.1 M phosphate buffer, pH 8.0
75
-
40% loss of activity within 30 min incubation, 0.1 M phosphate buffer, pH 8.0
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
glycerol, pyridoxal-5'-phosphate, dithiothreitol and EDTA do not increase the stability of the protein after freezing
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 10 mM phosphate buffer, pH 7, 0.01 mM pyridoxal phosphate, 0.1 mM DTT, 0.1 mM EDTA 50% glycerol, at least 2 months, no loss of activity
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
partial, does not bind to Blue-Sepharose affinity resin
-
partially by Sephadex G-100 and DEAE column chromatography
-
purified from Escherichia coli XL1-Blue cells harboring plasmid pKKyedO
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recombinant His6-tagged enzyme from Escherichia coli by nickel affinity chromatography
recombinant protein
-
Recombinant protein expressed with an N-terminal 6xHis-tag is purified under native/non-denaturing conditions using Ni-NTA Superflow resin.
-
The recombinant protein expressed with an N-terminal 6xHis-tag is purified under native/non-denaturing conditions using Ni-NTA Superflow resin
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloning into the pET30a (+) vector at the EcoRV/HindIII sites, all single and double mutants are constructed using a Phusion Site Directed Mutagenesis Kit.
-
epression of His6-tagged enzyme in Escherichia coli
expression in Escherichia coli
-
Expression of all recombinant proteins and mutants is carried out in Escherichia coli BL21. Altering of only two amino acid residues within the predicted active site served to change the enzyme from D-cysteine desulfhydrase to ACC deaminase.
overexpressed in host strain, enables Escherichia coli to utilize D-cysteine as sole sulfur source
-
semi-quantitative reverse transcription-PCR analysis
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
H2S fumigation stimulates the expression of drought associated genes. The enzyme is strongly induced by drought stress, with a maximum accumulation after 6 h, overview
the enzyme expression levels gradually increases in an age-dependent manner
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
E295S
-
By site-directed mutagenesis. The Pseudomonas putida UW4 single mutant is constructed using pET30a (+) with the full-length ACC deaminase as the template
E295S/L322T
-
The double mutant is constructed using the E295S mutant as the template.
E295S
-
By site-directed mutagenesis. The Pseudomonas putida UW4 single mutant is constructed using pET30a (+) with the full-length ACC deaminase as the template
-
E295S/L322T
-
The double mutant is constructed using the E295S mutant as the template.
-
H80Q
site-directed mutagenesis, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme
Q77H
site-directed mutagenesis, the mutant is inactive with respect to D-Cys, but retains significant 30% activity with beta-chloro-D-alanine
S78A
site-directed mutagenesis, the mutant is inactive with respect to D-Cys, but retains significant 20% activity with beta-chloro-D-alanine
T288E
site-directed mutagenesis, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme
T315L/T288E
site-directed mutagenesis, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme
Y261F
site-directed mutagenesis, the mutant shows altered substrate specificity and activity compared to the wild-type enzyme
Y287F
site-directed mutagenesis, inactive mutant with all substrates tested
S358E
By site-directed mutagenesis. The single mutants of the tomato enzyme are constructed using pET30 Xa/LIC with the full-length putative ACC deaminase as the template
S358E/T386L
The double mutant is constructed using an additional round of mutagenesis and with the S358E single mutant as the template
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