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Literature summary extracted from

  • Blanche, F.; Couder, M.; Debussche, L.; Thibaut, D.; Cameron, B.; Cruzet, J.
    Biosynthesis of vitamin B12: stepwise amidation of carboxyl groups b, d, e, and g of cobyrinic acid a,c-diamide is catalyzed by one enzyme in Pseudomonas denitrificans (1991), J. Bacteriol., 173, 6046-6051.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
6.3.5.9
-
 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 cobQ gene Pseudomonas denitrificans (nom. rej.)

General Stability

EC Number General Stability Organism
6.3.5.9 glycerol and DTT are necessary to stabilize the enzyme during purification Pseudomonas denitrificans (nom. rej.)
6.3.5.10 20% w/v and 1 mM dithiothreitol are necessary to stabilize enzyme during purification Pseudomonas denitrificans (nom. rej.)

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.3.5.9 0.00104
-
 adenosyl-cobyrinic acid a,c-diamide pH 7.5, 20°C Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.00121
-
 adenosyl-cobyrinic acid tetraamide pH 7.5, 20°C Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.00122
-
 adenosyl-cobyrinic acid triamide pH 7.5, 20°C Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.00125
-
 adenosyl-cobyrinic acid pentaamide pH 7.5, 20°C Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.011
-
 ATP pH 7.5, 20°C, adenosyl-cobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.033
-
 L-glutamine pH 7.5, 20°C, adenosyl-cobyrinic acid triamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.035
-
 ATP pH 7.5, 20°C, adenosyl-cobyrinic acid triamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.045
-
 L-glutamine pH 7.5, 20°C, adenosyl-cobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.049
-
 L-glutamine pH 7.5, 20°C, adenosyl-cobyrinic acid pentaamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.064
-
 L-glutamine pH 7.5, 20°C, adenosyl-cobyrinic acid tetraamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.088
-
 ATP pH 7.5, 20°C, adenosyl-cobyrinic acid pentaamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.12
-
 ATP pH 7.5, 20°C, adenosyl-cobyrinic acid tetraamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 20
-
 ammonia pH 7.5, 20°C Pseudomonas denitrificans (nom. rej.)
6.3.5.10 additional information
-
 additional information
-
 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.00104
-
 adenosylcobyrinic acid a,c-diamide pH 7.5 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.00121
-
 adenosylcobyrinic acid tetraamide pH 7.5 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.00122
-
 adenosylcobyrinic acid triamide pH 7.5 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.00125
-
 adenosylcobyrinic acid pentaamide pH 7.5 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.011
-
 ATP pH 7.5, adenosylcobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.033
-
 L-glutamine pH 7.5, adenosylcobyrinic acid triamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.035
-
 ATP pH 7.5, adenosylcobyrinic acid triamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.045
-
 L-glutamine pH 7.5, adenosylcobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.049
-
 L-glutamine pH 7.5, adenosylcobyrinic acid pentaamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.064
-
 L-glutamine pH 7.5, adenosylcobyrinic acid tetraamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.088
-
 ATP pH 7.5, adenosylcobyrinic acid pentaamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.12
-
 ATP pH 7.5, adenosylcobyrinic acid tetraamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 20.4
-
 ammonia pH 7.5, adenosylcobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
6.3.5.9 Mg2+ amidation reaction is Mg2+ dependent Pseudomonas denitrificans (nom. rej.)
6.3.5.10 Mg2+ Mg2+-dependent Pseudomonas denitrificans (nom. rej.)

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
6.3.5.9 57000
-
 2 * 57000, homodimer, SDS-PAGE Pseudomonas denitrificans (nom. rej.)
6.3.5.9 97300
-
 gel filtration Pseudomonas denitrificans (nom. rej.)
6.3.5.10 52000
-
 predicted from DNA sequence of the cobQ gene Pseudomonas denitrificans (nom. rej.)
6.3.5.10 57000
-
 2 * 57000, SDS-PAGE Pseudomonas denitrificans (nom. rej.)
6.3.5.10 97300
-
 gel filtration Pseudomonas denitrificans (nom. rej.)

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
6.3.5.9 5'-deoxy-5'-adenosyl-cobyrinic acid + 2 ATP + 2 glutamine + 2 H2O Pseudomonas denitrificans (nom. rej.) cobalamin biosynthetic pathway 5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + 2 ADP + 2 phosphate + 2 glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O Pseudomonas denitrificans (nom. rej.) biosynthesis of vitamin B12, cobalamin biosynthetic pathway ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O Pseudomonas denitrificans (nom. rej.) SC510 RifT biosynthesis of vitamin B12, cobalamin biosynthetic pathway ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
6.3.5.9 Pseudomonas denitrificans (nom. rej.)
-
-
-
6.3.5.9 Pseudomonas denitrificans (nom. rej.) SC510 RifT
-
-
-
6.3.5.10 Pseudomonas denitrificans (nom. rej.)
-
-
-
6.3.5.10 Pseudomonas denitrificans (nom. rej.)
-
 recombinant strain SC510 Rifr(pXL617)
-

Oxidation Stability

EC Number Oxidation Stability Organism
6.3.5.10 sensitive to oxidation Pseudomonas denitrificans (nom. rej.)

Purification (Commentary)

EC Number Purification (Comment) Organism
6.3.5.9
-
 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 45fold Pseudomonas denitrificans (nom. rej.)

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
6.3.5.9 0.068
-
 adenosyl-cobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0.083
-
 adenosyl-cobyrinic acid pentaamide as substrate Pseudomonas denitrificans (nom. rej.)
6.3.5.10 0.083
-
 pH 7.5, adenosylcobyrinic acid a,c-diamide as substrate Pseudomonas denitrificans (nom. rej.)

Storage Stability

EC Number Storage Stability Organism
6.3.5.9 -20°C, 0.1 M Tris hydrochloride, pH 7.5, 100% activity is lost after 48 h Pseudomonas denitrificans (nom. rej.)
6.3.5.9 -20°C, enzyme can be stored in the eluate from purification step v without detectable loss of activity Pseudomonas denitrificans (nom. rej.)
6.3.5.9 0°C, 0.1 M Tris hydrochloride, pH 7.5, 100% activity is lost after 48 h Pseudomonas denitrificans (nom. rej.)
6.3.5.10 -20°C or 0°C, purified enzyme, 0.1 M Tris hydrochloride, pH 7.5, 48 h, 100% loss of activity Pseudomonas denitrificans (nom. rej.)
6.3.5.10 -20°C, purified enzyme, 50 mM Tris hydrochloride, pH 7.5, 0.1 M sodium chloride, 1 mM dithiothreitol, 20% glycerol, several weeks, stable Pseudomonas denitrificans (nom. rej.)

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
6.3.5.9 5'-deoxy-5'-adenosyl-cobyrinic acid + 2 ATP + 2 glutamine + 2 H2O
-
 Pseudomonas denitrificans (nom. rej.) 5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + 2 ADP + 2 phosphate + 2 glutamate
-
 ?
6.3.5.9 5'-deoxy-5'-adenosyl-cobyrinic acid + 2 ATP + 2 glutamine + 2 H2O cobalamin biosynthetic pathway Pseudomonas denitrificans (nom. rej.) 5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + 2 ADP + 2 phosphate + 2 glutamate
-
 ?
6.3.5.9 5'-deoxy-5'-adenosyl-cobyrinic acid + ATP + ammonia + H2O
-
 Pseudomonas denitrificans (nom. rej.) 5'-deoxy-5'-adenosyl-cobyrinic acid ac-diamide + ADP + phosphate + ?
-
 ?
6.3.5.9 adenosyl-cobyrinic acid pentaamide + ATP + glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) ADP + glutamate + phosphate + ?
-
 ?
6.3.5.9 adenosyl-cobyrinic acid tetraamide + ATP + glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) ADP + glutamate + phosphate + ?
-
 ?
6.3.5.9 adenosyl-cobyrinic acid triamide + ATP + glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) ADP + glutamate + phosphate + ?
-
 ?
6.3.5.9 ATP + adenosyl-cobyrinic acid a,c-diamide + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) ADP + phosphate + ?
-
 ?
6.3.5.9 additional information adenosyl-cobyric acid and hydrogenobyrinic acid a,c-diamide are no substrates Pseudomonas denitrificans (nom. rej.) ?
-
 ?
6.3.5.10 adenosylcobyrinic acid a,c-diamide + ATP + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) adenosylcobyric acid + ADP + phosphate + L-glutamate
-
 ?
6.3.5.10 adenosylcobyrinic acid a,c-diamide + ATP + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) SC510 RifT adenosylcobyric acid + ADP + phosphate + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + ammonia + H2O ammonia can replace L-glutamine as amide group donor, but L-glutamine is preferred Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + ?
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O biosynthesis of vitamin B12, cobalamin biosynthetic pathway Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O enzyme catalyzes the stepwise amidation of carboxyl groups b, d, e, and g of cobyrinic acid a,c-diamide, four-step amidation sequence from cobyrinic acid a,c-diamide to cobyric acid via the formation of corbyrinic acid triamide, tetraamide, and pentaamide intermediates, the amidations are carried out in a specific order, enzyme is specific to coenzyme forms of substrates, no amidation of the carboxyl group at position f, L-glutamine is the preferred amide group donor Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O biosynthesis of vitamin B12, cobalamin biosynthetic pathway Pseudomonas denitrificans (nom. rej.) SC510 RifT ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid a,c-diamide + L-glutamine + H2O enzyme catalyzes the stepwise amidation of carboxyl groups b, d, e, and g of cobyrinic acid a,c-diamide, four-step amidation sequence from cobyrinic acid a,c-diamide to cobyric acid via the formation of corbyrinic acid triamide, tetraamide, and pentaamide intermediates, the amidations are carried out in a specific order, enzyme is specific to coenzyme forms of substrates, no amidation of the carboxyl group at position f, L-glutamine is the preferred amide group donor Pseudomonas denitrificans (nom. rej.) SC510 RifT ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid pentaamide + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid tetraamide + L-glutamine + H2O two-step amidation Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 ATP + adenosylcobyrinic acid triamide + L-glutamine + H2O three-step amidation Pseudomonas denitrificans (nom. rej.) ADP + phosphate + adenosylcobyric acid + L-glutamate
-
 ?
6.3.5.10 cobinic acid a,c-diamide + ATP + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) cobinamide + ADP + phosphate + L-glutamate
-
 ?
6.3.5.10 cobinic acid a,c-diamide + ATP + L-glutamine + H2O
-
 Pseudomonas denitrificans (nom. rej.) SC510 RifT cobinamide + ADP + phosphate + L-glutamate
-
 ?
6.3.5.10 additional information no amidation of adenosylcobyric acid, hydrogenobyrinic acid a,c-diamide, monocyano or diaqua form of cobyrinic acid a,c-diamide Pseudomonas denitrificans (nom. rej.) ?
-
 ?
6.3.5.10 additional information no amidation of adenosylcobyric acid, hydrogenobyrinic acid a,c-diamide, monocyano or diaqua form of cobyrinic acid a,c-diamide Pseudomonas denitrificans (nom. rej.) SC510 RifT ?
-
 ?

Subunits

EC Number Subunits Comment Organism
6.3.5.9 dimer 2 * 57000, homodimer, SDS-PAGE Pseudomonas denitrificans (nom. rej.)
6.3.5.10 dimer 2 * 57000, SDS-PAGE Pseudomonas denitrificans (nom. rej.)
6.3.5.10 homodimer 2 * 57000, SDS-PAGE Pseudomonas denitrificans (nom. rej.)

Synonyms

EC Number Synonyms Comment Organism
6.3.5.9 cobyric acid synthase
-
 Pseudomonas denitrificans (nom. rej.)
6.3.5.10 CobQ
-
 Pseudomonas denitrificans (nom. rej.)

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
6.3.5.9 7.5
-
 broad optimum around Pseudomonas denitrificans (nom. rej.)
6.3.5.10 7.5
-
 broad pH-optimum around Pseudomonas denitrificans (nom. rej.)

Cofactor

EC Number Cofactor Comment Organism Structure
6.3.5.10 ATP ATP-dependent Pseudomonas denitrificans (nom. rej.)

pI Value

EC Number Organism Comment pI Value Maximum pI Value
6.3.5.9 Pseudomonas denitrificans (nom. rej.)
-
 5.9 5.6
6.3.5.10 Pseudomonas denitrificans (nom. rej.) isoelectric focusing
-
 6