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Literature summary for 5.4.3.5 extracted from

  • Makins, C.; Pickering, A.V.; Mariani, C.; Wolthers, K.R.
    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.
    View publication on PubMed

Protein Variants

Protein Variants Comment Organism
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
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
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
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

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information pre-steady-state and stedy-state kinetics of wild-type and mutant enzymes Acetoanaerobium sticklandii
0.03
-
D-ornithine recombinant His-tagged mutant E338A, pH 7.5, 30°C Acetoanaerobium sticklandii
0.043
-
D-ornithine recombinant His-tagged mutant E338D, pH 7.5, 30°C Acetoanaerobium sticklandii
0.061
-
D-ornithine recombinant His-tagged mutant E338Q, pH 7.5, 30°C Acetoanaerobium sticklandii
0.19
-
D-ornithine recombinant His-tagged wild-type enzyme, pH 7.5, 30°C Acetoanaerobium sticklandii

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
D-ornithine Acetoanaerobium sticklandii
-
(2R,4S)-2,4-diaminopentanoate
-
?
D-ornithine Acetoanaerobium sticklandii DSM 519
-
(2R,4S)-2,4-diaminopentanoate
-
?

Organism

Organism UniProt Comment Textmining
Acetoanaerobium sticklandii E3PY95 beta-subunit
-
Acetoanaerobium sticklandii E3PY96 alpha-subunit
-
Acetoanaerobium sticklandii DSM 519 E3PY95 beta-subunit
-
Acetoanaerobium sticklandii DSM 519 E3PY96 alpha-subunit
-

Reaction

Reaction Comment Organism Reaction ID
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
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

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-ornithine
-
Acetoanaerobium sticklandii (2R,4S)-2,4-diaminopentanoate
-
?
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
-
?
D-ornithine
-
Acetoanaerobium sticklandii DSM 519 (2R,4S)-2,4-diaminopentanoate
-
?
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
-
?

Synonyms

Synonyms Comment Organism
OAM
-
Acetoanaerobium sticklandii
ornithine 4,5-aminomutase
-
Acetoanaerobium sticklandii

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
assay at Acetoanaerobium sticklandii

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.032
-
D-ornithine recombinant His-tagged mutant E338D, pH 7.5, 30°C Acetoanaerobium sticklandii
2.9
-
D-ornithine recombinant His-tagged wild-type enzyme, pH 7.5, 30°C Acetoanaerobium sticklandii
4.3
-
D-ornithine recombinant His-tagged mutant E338Q, pH 7.5, 30°C Acetoanaerobium sticklandii
7.6
-
D-ornithine recombinant His-tagged mutant E338A, pH 7.5, 30°C Acetoanaerobium sticklandii

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Acetoanaerobium sticklandii

Cofactor

Cofactor Comment Organism Structure
adenosylcobalamin binding analysis with recombinant wild-type and mutant enzymes, overview Acetoanaerobium sticklandii

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.07
-
D-ornithine recombinant His-tagged mutant E338Q, pH 7.5, 30°C Acetoanaerobium sticklandii
0.25
-
D-ornithine recombinant His-tagged mutant E338A, pH 7.5, 30°C Acetoanaerobium sticklandii
0.75
-
D-ornithine recombinant His-tagged mutant E338D, pH 7.5, 30°C Acetoanaerobium sticklandii
15.2
-
D-ornithine recombinant His-tagged wild-type enzyme, pH 7.5, 30°C Acetoanaerobium sticklandii