Information on EC 6.3.5.9 - hydrogenobyrinic acid a,c-diamide synthase (glutamine-hydrolysing)

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
6.3.5.9
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RECOMMENDED NAME
GeneOntology No.
hydrogenobyrinic acid a,c-diamide synthase (glutamine-hydrolysing)
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
2 ATP + hydrogenobyrinic acid + 2 L-glutamine + 2 H2O = 2 ADP + 2 phosphate + hydrogenobyrinic acid a,c-diamide + 2 L-glutamate
show the reaction diagram
This step in the aerobic biosynthesis of cobalamin generates hydrogenobyrinic acid a,c-diamide, the substrate required by EC 6.6.1.2, cobaltochelatase, which adds cobalt to the macrocycle
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PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
cob(II)yrinate a,c-diamide biosynthesis II (late cobalt incorporation)
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Metabolic pathways
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Porphyrin and chlorophyll metabolism
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vitamin B12 metabolism
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SYSTEMATIC NAME
IUBMB Comments
hydrogenobyrinic-acid:L-glutamine amido-ligase (AMP-forming)
This step in the aerobic biosynthesis of cobalamin generates hydrogenobyrinic acid a,c-diamide, the substrate required by EC 6.6.1.2, cobaltochelatase, which adds cobalt to the macrocycle.
CAS REGISTRY NUMBER
COMMENTARY hide
132053-22-6
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
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Manually annotated by BRENDA team
strain SC510
UniProt
Manually annotated by BRENDA team
strain SC510 RifT
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Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
5'-deoxy-5'-adenosyl-cobyrinic acid + 2 ATP + 2 glutamine + 2 H2O
5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + 2 ADP + 2 phosphate + 2 glutamate
show the reaction diagram
5'-deoxy-5'-adenosyl-cobyrinic acid + ATP + ammonia + H2O
5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + ADP + phosphate + ?
show the reaction diagram
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-
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?
adenosyl-cobyrinic acid pentaamide + ATP + glutamine + H2O
ADP + glutamate + phosphate + ?
show the reaction diagram
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?
adenosyl-cobyrinic acid tetraamide + ATP + glutamine + H2O
ADP + glutamate + phosphate + ?
show the reaction diagram
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?
adenosyl-cobyrinic acid triamide + ATP + glutamine + H2O
ADP + glutamate + phosphate + ?
show the reaction diagram
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?
alpha-ribazole + phosphate
N1-(5-phospho-alpha-D-ribosyl)-5,6-dimethylbenzimidazole + H2O
show the reaction diagram
ATP + (aq)2cobyrinic acid + glutamine + H2O
ADP + phosphate + ?
show the reaction diagram
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?
ATP + (CN,aq)cobyrinic acid + glutamine + H2O
ADP + phosphate + c-monoamide + ?
show the reaction diagram
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?
ATP + (CN,aq)cobyrinic acid -monoamide + glutamine + H2O
ADP + phosphate + ?
show the reaction diagram
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?
ATP + cobyrinic acid + ammonia + H2O
ADP + phosphate + ?
show the reaction diagram
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?
ATP + cobyrinic acid + L-glutamine + H2O
ADP + phosphate + cobyrinic acid a,c-diamide + L-glutamate
show the reaction diagram
ATP + hydrogenobyrinic acid + L-glutamine + H2O
ADP + phosphate + hydrogenobyrinic acid a,c-diamide + L-glutamate
show the reaction diagram
ATP + hydrogenobyrinic acid c-monoamide + L-glutamine + H2O
ADP + phosphate + ?
show the reaction diagram
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?
cobyrinic acid + ATP + glutamine
cobyrinic acid a,c-diamide + ADP + phosphate + glutamate
show the reaction diagram
cobyrinic acid + ATP + NH4+
cobyrinic acid a,c-diamide + ADP + phosphate
show the reaction diagram
cobyrinic acid c-monoamide + ATP + glutamine
cobyrinic acid a,c-diamide + ADP + phosphate + glutamate
show the reaction diagram
cobyrinic acid c-monoamide + ATP + NH4+
cobyrinic acid a,c-diamide + ADP + phosphate
show the reaction diagram
DMB + nicotinic acid mononucleotide
DMB-ribose-5'-phosphate
show the reaction diagram
additional information
?
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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
5'-deoxy-5'-adenosyl-cobyrinic acid + 2 ATP + 2 glutamine + 2 H2O
5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + 2 ADP + 2 phosphate + 2 glutamate
show the reaction diagram
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cobalamin biosynthetic pathway
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?
ATP + cobyrinic acid + L-glutamine + H2O
ADP + phosphate + cobyrinic acid a,c-diamide + L-glutamate
show the reaction diagram
P21632
cobalamin pathway from cobyrinic acid
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?
ATP + hydrogenobyrinic acid + L-glutamine + H2O
ADP + phosphate + hydrogenobyrinic acid a,c-diamide + L-glutamate
show the reaction diagram
additional information
?
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.25
(aq)2cobyrinic acid
pH 7.6, 30C
0.16
(CN,aq)cobyrinic acid
pH 7.6, 30C
0.071
(CN,aq)cobyrinic acid c-monoamide
pH 7.6, 30C
0.00104
adenosyl-cobyrinic acid a,c-diamide
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pH 7.5, 20C
0.00125
adenosyl-cobyrinic acid pentaamide
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pH 7.5, 20C
0.00121
adenosyl-cobyrinic acid tetraamide
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pH 7.5, 20C
0.00122
adenosyl-cobyrinic acid triamide
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pH 7.5, 20C
0.012 - 20
ammonia
0.0027 - 0.12
ATP
0.00074
cobyrinic acid
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pH 7.7, wild-type enzyme
0.0003 - 0.011
cobyrinic acid c-monoamide
0.0203 - 0.053
glutamine
0.00041
Hydrogenobyrinic acid
pH 7.6, 30C
0.00021
hydrogenobyrinic acid c-monoamide
pH 7.6, 30C
0.033 - 0.064
L-glutamine
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.13
ATP
Salmonella enterica subsp. enterica serovar Typhimurium
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pH 7.7, wild-type enzyme
0.16
cobyrinic acid
Salmonella enterica subsp. enterica serovar Typhimurium
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pH 7.7, wild-type enzyme
0.008 - 0.2
cobyrinic acid c-monoamide
0.16
glutamine
Salmonella enterica subsp. enterica serovar Typhimurium
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pH 7.7, wild-type enzyme
0.18
NH4+
Salmonella enterica subsp. enterica serovar Typhimurium
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pH 7.7, wild-type enzyme
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.068
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adenosyl-cobyrinic acid a,c-diamide as substrate
0.083
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adenosyl-cobyrinic acid pentaamide as substrate
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5
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broad optimum around
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
26300
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protein predicted from nucleotide sequence
57000
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2 * 57000, homodimer, SDS-PAGE
86000
gel filtration
97300
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
glycerol and DTT are necessary to stabilize the enzyme during purification
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, 0.1 M Tris hydrochloride, pH 7.5, 100% activity is lost after 48 h
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-20C, enzyme can be stored in the eluate from purification step v without detectable loss of activity
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0C, 0.1 M Tris hydrochloride, pH 7.5, 100% activity is lost after 48 h
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned, sequenced and overexpressed
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expression of wild-type and mutant enzymes in Escherichia coli
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identification of the structural gene
plasmid pXL 191 is transferre from Escherichia coli MC1060 to Pseudomonas denitrificans SC510 RifT through conjugation
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D45N
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less than 0.2% of amidation activity with glutamine and with NH4+, compared to wild-type values
D48N
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less than 0.3% of amidation activity with glutamine and with NH4+, compared to wild-type values
D97N
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32.4% of wild-type amidation activity with glutamine, 16.5% of wild-type amidation activity with NH4+
E90Q
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less than 0.02% of amidation activity with glutamine and with NH4+, compared to wild-type values
L47A
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10.6% of wild-type amidation activity with glutamine, 44% of wild-type amidation activity with NH4+, mutation specifically decreases the affinity of the enzyme to the cobyrinic acid c-monoamide
Y46A
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3.8% of wild-type amidation activity with glutamine, 2% of wild-type amidation activity with NH4+, mutation specifically decreases the affinity of the enzyme to the cobyrinic acid c-monoamide
D45N
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less than 0.2% of amidation activity with glutamine and with NH4+, compared to wild-type values
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D48N
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less than 0.3% of amidation activity with glutamine and with NH4+, compared to wild-type values
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D97N
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32.4% of wild-type amidation activity with glutamine, 16.5% of wild-type amidation activity with NH4+
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E90Q
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less than 0.02% of amidation activity with glutamine and with NH4+, compared to wild-type values
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Y46A
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3.8% of wild-type amidation activity with glutamine, 2% of wild-type amidation activity with NH4+, mutation specifically decreases the affinity of the enzyme to the cobyrinic acid c-monoamide
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Show AA Sequence (2059 entries)
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