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Literature summary extracted from
- Mutagenesis of a conserved glutamate reveals the contribution of electrostatic energy to adenosylcobalamin co-C bond homolysis in ornithine 4,5-aminomutase and methylmalonyl-CoA mutase (2013), Biochemistry, 52, 878-888.
Cloned(Commentary)
| EC Number | Cloned (Comment) | Organism |
|---|---|---|
| 5.4.99.2 | expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain Rosetta(DE3)pLysS cells | Homo sapiens |
Protein Variants
| EC Number | Protein Variants | Comment | Organism |
|---|---|---|---|
| 5.4.3.5 | E338A | site-directed mutagenesis, substrate binding of the mutant is unaffected, but kcat is reduced 670fold and catalytic efficiency 220fold compared to the wild-type enzyme. The rate of external aldimine formation in the mutant is similar to that of the wild-type enzyme, but it shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Acetoanaerobium sticklandii |
| 5.4.3.5 | E338D | site-directed mutagenesis, substrate binding of the mutant is unaffected, but kcat is reduced 380fold and catalytic efficiency 60fold compared to the wild-type enzyme. The rate of external aldimine formation in the mutant is similar to that of the wild-type enzyme, but it shows n detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Acetoanaerobium sticklandii |
| 5.4.3.5 | E338D | site-directed mutagenesis, substrate binding of the mutant is unaffected, but kcat is reduced 380fold and catalytic efficiency 60fold compared to the wild-type enzyme. The rate of external aldimine formation in the mutant is similar to that of the wild-type enzyme, but it shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Acetoanaerobium sticklandii |
| 5.4.3.5 | E338Q | site-directed mutagenesis, substrate binding of the mutant is unaffected, but kcat is reduced 90fold and catalytic efficiency 20fold compared to the wild-type enzyme. The rate of external aldimine formation in the mutant is similar to that of the wild-type enzyme, but it shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Acetoanaerobium sticklandii |
| 5.4.99.2 | E392A | site-directed mutagenesis, kcat is reduced 12fold compared to the wild-type enzyme. The mutant shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Homo sapiens |
| 5.4.99.2 | E392D | site-directed mutagenesis, kcat is reduced 330fold compared to the wild-type enzyme. The mutant shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Homo sapiens |
| 5.4.99.2 | E392Q | site-directed mutagenesis, kcat is reduced 16fold compared to the wild-type enzyme. The mutant shows no detectable adenosylcobalamin homolysis upon binding of the physiological substrate | Homo sapiens |
KM Value [mM]
| EC Number | KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 5.4.3.5 | additional information | - |
additional information | pre-steady-state and stedy-state kinetics of wild-type and mutant enzymes | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 0.03 | - |
D-ornithine | recombinant His-tagged mutant E338A, pH 7.5, 30°C | Acetoanaerobium sticklandii |  |
| 5.4.3.5 | 0.043 | - |
D-ornithine | recombinant His-tagged mutant E338D, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 0.061 | - |
D-ornithine | recombinant His-tagged mutant E338Q, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 0.19 | - |
D-ornithine | recombinant His-tagged wild-type enzyme, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.99.2 | additional information | - |
additional information | pre-steady-state and steady-state kinetics of wild-type and mutant enzymes | Homo sapiens |
Localization
| EC Number | Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|---|
| 5.4.99.2 | mitochondrion | - |
Homo sapiens | 5739 | - |
Natural Substrates/ Products (Substrates)
| EC Number | Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 5.4.3.5 | D-ornithine | Acetoanaerobium sticklandii | - |
(2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.3.5 | D-ornithine | Acetoanaerobium sticklandii DSM 519 | - |
(2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.99.2 | (R)-methylmalonyl-CoA | Homo sapiens | - |
succinyl-CoA | - |
? | |
Organism
| EC Number | Organism | UniProt | Comment | Textmining |
|---|---|---|---|---|
| 5.4.3.5 | Acetoanaerobium sticklandii | E3PY95 | beta-subunit | - |
| 5.4.3.5 | Acetoanaerobium sticklandii | E3PY96 | alpha-subunit | - |
| 5.4.3.5 | Acetoanaerobium sticklandii DSM 519 | E3PY95 | beta-subunit | - |
| 5.4.3.5 | Acetoanaerobium sticklandii DSM 519 | E3PY96 | alpha-subunit | - |
| 5.4.99.2 | Homo sapiens | P22033 | - |
- |
Purification (Commentary)
| EC Number | Purification (Comment) | Organism |
|---|---|---|
| 5.4.99.2 | recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain Rosetta(DE3)pLysS cells by nickel affinity and anion exchange chromatography | Homo sapiens |
Reaction
| EC Number | Reaction | Comment | Organism | Reaction ID |
|---|---|---|---|---|
| 5.4.3.5 | D-ornithine = (2R,4S)-2,4-diaminopentanoate | mechanism, overview, a gradual weakening of the electrostatic energy between the protein and the ribose leads to a progressive increase in the activation energy barrier for adenosylcobalamin Co-C bond homolysis, key role for the conserved polar glutamate residue in controlling the initial generation of radical species | Acetoanaerobium sticklandii | |
| 5.4.3.5 | D-ornithine = (2R,4S)-2,4-diaminopentanoate | mechanism, overview, a gradual weakening of the electrostatic energy between the protein and the ribose leads to a progressive increase in the activation energy barrier for adenosylcobalamin Co?C bond homolysis, key role for the conserved polar glutamate residue in controlling the initial generation of radical species | Acetoanaerobium sticklandii | |
| 5.4.99.2 | (R)-methylmalonyl-CoA = succinyl-CoA | mechanism, overview, a gradual weakening of the electrostatic energy between the protein and the ribose leads to a progressive increase in the activation energy barrier for Co-C bond homolysis, key role for the conserved polar glutamate residue in controlling the initial generation of radical species | Homo sapiens |
Substrates and Products (Substrate)
| EC Number | Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|---|
| 5.4.3.5 | D-ornithine | - |
Acetoanaerobium sticklandii | (2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.3.5 | D-ornithine | binding of substrate to the enzyme leads to the formation of an electrostatic interaction between a conserved glutamate side chain and the adenosyl ribose of the adenosylcobalamin cofactor. Residue Glu338 is involved in adenosylcobalamin Co-C bond labilization and catalysis | Acetoanaerobium sticklandii | (2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.3.5 | D-ornithine | - |
Acetoanaerobium sticklandii DSM 519 | (2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.3.5 | D-ornithine | binding of substrate to the enzyme leads to the formation of an electrostatic interaction between a conserved glutamate side chain and the adenosyl ribose of the adenosylcobalamin cofactor. Residue Glu338 is involved in adenosylcobalamin Co-C bond labilization and catalysis | Acetoanaerobium sticklandii DSM 519 | (2R,4S)-2,4-diaminopentanoate | - |
? | |
| 5.4.99.2 | (R)-methylmalonyl-CoA | - |
Homo sapiens | succinyl-CoA | - |
? | |
| 5.4.99.2 | (R)-methylmalonyl-CoA | binding of substrate to the enzyme leads to the formation of an electrostatic interaction between a conserved glutamate side chain and the adenosyl ribose of the adenosylcobalamin cofactor. Residue Glu392 is involved in adenosylcobalamin Co-C bond labilization and catalysis | Homo sapiens | succinyl-CoA | - |
? | |
Synonyms
| EC Number | Synonyms | Comment | Organism |
|---|---|---|---|
| 5.4.3.5 | OAM | - |
Acetoanaerobium sticklandii |
| 5.4.3.5 | ornithine 4,5-aminomutase | - |
Acetoanaerobium sticklandii |
| 5.4.99.2 | MCM | - |
Homo sapiens |
Temperature Optimum [°C]
| EC Number | Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|---|
| 5.4.3.5 | 30 | - |
assay at | Acetoanaerobium sticklandii |
| 5.4.99.2 | 25 | - |
assay at | Homo sapiens |
Turnover Number [1/s]
| EC Number | Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 5.4.3.5 | 0.032 | - |
D-ornithine | recombinant His-tagged mutant E338D, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 2.9 | - |
D-ornithine | recombinant His-tagged wild-type enzyme, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 4.3 | - |
D-ornithine | recombinant His-tagged mutant E338Q, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 7.6 | - |
D-ornithine | recombinant His-tagged mutant E338A, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.99.2 | 0.012 | - |
(R)-methylmalonyl-CoA | recombinant His-tagged mutant E392D, pH 8.5, 25°C | Homo sapiens | |
| 5.4.99.2 | 0.25 | - |
(R)-methylmalonyl-CoA | recombinant His-tagged mutant E392Q, pH 8.5, 25°C | Homo sapiens | |
| 5.4.99.2 | 0.32 | - |
(R)-methylmalonyl-CoA | recombinant His-tagged mutant E392A, pH 8.5, 25°C | Homo sapiens | |
| 5.4.99.2 | 3.95 | - |
(R)-methylmalonyl-CoA | recombinant His-tagged wild-type enzyme, pH 8.5, 25°C | Homo sapiens | |
pH Optimum
| EC Number | pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|---|
| 5.4.3.5 | 7.5 | - |
assay at | Acetoanaerobium sticklandii |
| 5.4.99.2 | 8.5 | - |
assay at | Homo sapiens |
Cofactor
| EC Number | Cofactor | Comment | Organism | Structure |
|---|---|---|---|---|
| 5.4.3.5 | adenosylcobalamin | binding analysis with recombinant wild-type and mutant enzymes, overview | Acetoanaerobium sticklandii | |
| 5.4.99.2 | adenosylcobalamin | binding analysis with recombinant wild-type and mutant enzymes, overview | Homo sapiens | |
kcat/KM [mM/s]
| EC Number | kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
|---|---|---|---|---|---|---|
| 5.4.3.5 | 0.07 | - |
D-ornithine | recombinant His-tagged mutant E338Q, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 0.25 | - |
D-ornithine | recombinant His-tagged mutant E338A, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 0.75 | - |
D-ornithine | recombinant His-tagged mutant E338D, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
| 5.4.3.5 | 15.2 | - |
D-ornithine | recombinant His-tagged wild-type enzyme, pH 7.5, 30°C | Acetoanaerobium sticklandii | |
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