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Information on EC 4.2.1.103 - cyclohexyl-isocyanide hydratase and Organism(s) Pseudomonas putida and UniProt Accession Q8G9F9

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EC Tree
     4 Lyases
         4.2 Carbon-oxygen lyases
             4.2.1 Hydro-lyases
                4.2.1.103 cyclohexyl-isocyanide hydratase
IUBMB Comments
The enzyme from Pseudomonas putida strain N19-2 can also catalyse the hydration of other isonitriles to the corresponding N-substituted formamides. The enzyme has no metal requirements.
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This record set is specific for:
Pseudomonas putida
UNIPROT: Q8G9F9
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The taxonomic range for the selected organisms is: Pseudomonas putida
The expected taxonomic range for this enzyme is: Bacteria, Archaea
Synonyms
isonitrile hydratase, isocyanide hydratase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
isonitrile hydratase
-
cyclohexyl isocyanide hydratase
-
-
-
-
isocyanide hydratase
-
-
-
-
isonitrile hydratase
-
-
-
-
N-cyclohexylformamide hydro-lyase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
N-cyclohexylformamide = cyclohexyl isocyanide + H2O
show the reaction diagram
the enzyme from Pseudomonas putida strain N19-2 can also catalyse the hydration of other isonitriles to the corresponding N-substituted formamides, active site Cys101 and T102 are essential for activity, while Glu79 and Glu81 are not
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
addition
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
N-cyclohexylformamide hydro-lyase (cyclohexyl-isocyanide-forming)
The enzyme from Pseudomonas putida strain N19-2 can also catalyse the hydration of other isonitriles to the corresponding N-substituted formamides. The enzyme has no metal requirements.
CAS REGISTRY NUMBER
COMMENTARY hide
358974-06-8
-
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
cyclohexyl isocyanide + H2O
N-cyclohexylformamide
show the reaction diagram
-
-
?
benzyl isocyanide
N-phenylformamide
show the reaction diagram
-
224% of the activity with cyclohexyl isocyanide
-
-
?
cyclohexyl isocyanide + H2O
N-cyclohexylformamide
show the reaction diagram
isocyanomethyl phosphonic acid diethyl ether + H2O
formylaminomethyl-phosphonic acid diethyl ester
show the reaction diagram
-
3% of the activity with cyclohexyl isocyanide
-
-
?
additional information
?
-
the enzyme can also catalyse the hydration of other isonitriles to the corresponding N-substituted formamides
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
cyclohexyl isocyanide + H2O
N-cyclohexylformamide
show the reaction diagram
-
enzyme is involved in nitrogen-carbon triple bond cleavage
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
no metal requirements
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
AgNO3
-
1 mM, 99% inhibition
ammonium persulfate
-
1 mM, 23% inhibition
CdCl2
-
1 mM, 87% inhibition
CoCl2
-
1 mM, 98% inhibition
CuSO4
-
1 mM, 98% inhibition
Diethyldithiocarbamic acid sodium salt
-
1 mM, 20% inhibition
FeSO4
-
1 mM, 50% inhibition
H2O2
-
1 mM, 49% inhibition
HgCl2
-
1 mM, 99% inhibition
hydroxylamine
-
1 mM, 10% inhibition
iodoacetate
-
1 mM, 97% inhibition
KCN
-
1 mM, 26% inhibition
NEM
-
1 mM,51% inhibition
NiSO4
-
1 mM, 98% inhibition
PCMB
-
1 mM, 50% inhibition
phenylmethanesulfonyl fluoride
-
1 mM, 57% inhibition
SnCl2
-
1 mM, 16% inhibition
ZnCl2
-
1 mM, 34% inhibition
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
16.2
cyclohexyl isocyanide
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
1.4
purified recombinant mutant T102A
15.4
purified recombinant mutant E81Q
17.3
purified recombinant wild-type enzyme
18.7
purified recombinant mutant E79Q
additional information
-
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.5 - 9
-
pH 5.5: about 45% of maximal activity, potassium phosphate buffer, pH 9.0: about 30% of maximal activity, NH4OH/NH4Cl buffer
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20 - 45
-
20°C: about 55% of maximal activity, 45°C: about 55% of maximal activity
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
SwissProt
Manually annotated by BRENDA team
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
INHA_PSEPU
228
0
24213
Swiss-Prot
-
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
24211
x * 24211, amino acid sequence calculation
59000
-
2 * 59000, SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
x * 24211, amino acid sequence calculation
dimer
-
2 * 59000, SDS-PAGE
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C101A
site-directed mutagenesis, inactive mutant
E79Q
site-directed mutagenesis, slightly increased activity compared to the wild-type enzyme
E81Q
site-directed mutagenesis, slightly decreased activity compared to the wild-type enzyme
T102A
site-directed mutagenesis, highly reduced activity compared to the wild-type enzyme
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
10
-
30 min, 10% v/v glycerol, 3% loss of activity
20
-
30 min, 10% v/v glycerol, 11% loss of activity
30
-
30 min, 10% v/v glycerol, 16% loss of activity
35
-
30 min, 10% v/v glycerol, 22% loss of activity
40
-
30 min, 10% v/v glycerol, 31% loss of activity
45
-
30 min, 10% v/v glycerol, 47% loss of activity
50
-
30 min, 10% v/v glycerol, 67% loss of activity
55
-
30 min, 10% v/v glycerol, 81% loss of activity
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant wild-type and mutants from Escherichia coli
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene inhA, DNA and amino acid sequence determination and analysis, overexpression of wild-type and mutant enzyme in Escherichia coli
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Goda, M.; Hashimoto, Y.; Shimizu, S.; Kobayashi, M.
Discovery of a novel enzyme, isonitrile hydratase, involved in nitrogen-carbon triple bond cleavage
J. Biol. Chem.
276
23480-23485
2001
Pseudomonas putida, Pseudomonas putida N19-2
Manually annotated by BRENDA team
Goda, M.; Hashimoto, Y.; Takase, M.; Herai, S.; Iwahara, Y.; Higashibata, H.; Kobayashi, M.
Isonitrile hydratase from Pseudomonas putida N19-2. Cloning, sequencing, gene expression, and identification of its active acid residue
J. Biol. Chem.
277
45860-45865
2002
Pseudomonas putida (Q8G9F9), Pseudomonas putida N19-2 (Q8G9F9), Pseudomonas putida N19-2
Manually annotated by BRENDA team