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Literature summary extracted from

  • Komarova, N.V.; Golubev, I.V.; Khoronenkova, S.V.; Chubar, T.A.; Tishkov, V.I.
    Engineering of substrate specificity of D-amino acid oxidase from the yeast Trigonopsis variabilis: directed mutagenesis of Phe258 residue (2012), Biochemistry, 77, 1181-1189.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.4.3.3 expressed in Escherichia coli BL21(DE3) CodonPlus/pLysS cells Trigonopsis variabilis

Protein Variants

EC Number Protein Variants Comment Organism
1.4.3.3 F258A the improvement of catalytic efficiency with D-Tyr, D-Phe, and D-Leu for mutant enzyme F258A is 3.66, 11.7, and 1.5fold, respectively. The mutant is inactive with D-Ala, D-Ser, D-Lys, and D-Thr, while the activity with D-Tyr, D-Leu and especially with D-Phe significantly increases Trigonopsis variabilis
1.4.3.3 F258S the improvement of catalytic efficiency with D-Tyr, D-Phe, and D-Leu for mutant enzyme F258S is 1.7, 4.75, and 6.61fold, respectively. The mutant is inactive with D-Ala, D-Ser, D-Lys, and D-Thr, while the activity with D-Tyr, D-Leu and especially with D-Phe significantly increases Trigonopsis variabilis
1.4.3.3 F258Y the mutant is inactive with D-Val, D-Tyr, D-Ala, D-Ser, D-Lys, and D-Thr Trigonopsis variabilis

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.4.3.3 0.068
-
 D-Phe mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.087
-
 D-Tyr mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.134
-
 D-Phe mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.227
-
 D-Leu mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.37
-
 D-Phe wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.45
-
 D-Trp mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.45
-
 D-Tyr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.46
-
 D-Trp mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.46
-
 D-Met wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.49
-
 D-Trp wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.78
-
 D-Leu wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.84
-
 D-Tyr mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 1.19
-
 D-Leu mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 1.68
-
 D-Phe mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 1.81
-
 D-Leu mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 2.7
-
 D-Trp mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 2.9
-
 D-Met mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 3.3
-
 D-Met mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 4.2
-
 D-Met mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 11.1
-
 D-Thr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 14.4
-
 D-Val wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 16.7
-
 D-Ala wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 22
-
 D-Asn mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 22.6
-
 D-Asn wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 29
-
 D-Asn mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 29.3
-
 D-Lys wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 36.6
-
 D-Ser wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 41
-
 D-Val mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 46
-
 D-Val mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 50
-
 D-Asn Km above 50 mM, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.4.3.3 38000
-
 x * 38000, SDS-PAGE Trigonopsis variabilis

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.4.3.3 D-amino acid + H2O + O2 Trigonopsis variabilis
-
 2-oxocarboxylate + NH3 + H2O2
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.4.3.3 Trigonopsis variabilis
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.4.3.3 MonoQ column chromatography and Sephadex G-25 gel filtration Trigonopsis variabilis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.4.3.3 D-Ala + H2O + O2
-
 Trigonopsis variabilis pyruvate + NH3 + H2O2
-
 ?
1.4.3.3 D-amino acid + H2O + O2
-
 Trigonopsis variabilis 2-oxocarboxylate + NH3 + H2O2
-
 ?
1.4.3.3 D-Asn + H2O + O2
-
 Trigonopsis variabilis 2-oxosuccinamate + NH3 + H2O2
-
 ?
1.4.3.3 D-Leu + H2O + O2
-
 Trigonopsis variabilis 4-methyl-2-oxopentanoic acid + NH3 + H2O2
-
 ?
1.4.3.3 D-Lys + H2O + O2
-
 Trigonopsis variabilis 6-amino-2-oxohexanoic acid + NH3 + H2O2
-
 ?
1.4.3.3 D-Met + H2O + O2 best substrate Trigonopsis variabilis 4-methylthio-2-oxobutanoic acid + NH3 + H2O2
-
 ?
1.4.3.3 D-Phe + H2O + O2
-
 Trigonopsis variabilis phenylpyruvate + NH3 + H2O2
-
 ?
1.4.3.3 D-Ser + H2O + O2
-
 Trigonopsis variabilis 2-oxo-3-hydroxypropionic acid + NH3 + H2O2
-
 ?
1.4.3.3 D-Thr + H2O + O2
-
 Trigonopsis variabilis 2-oxo-3-hydroxybutyrate + NH3 + H2O2
-
 ?
1.4.3.3 D-Trp + H2O + O2
-
 Trigonopsis variabilis indol-3-pyruvate + NH3 + H2O2
-
 ?
1.4.3.3 D-Tyr + H2O + O2
-
 Trigonopsis variabilis 4-hydroxyphenylpyruvic acid + NH3 + H2O2
-
 ?
1.4.3.3 D-Val + H2O + O2
-
 Trigonopsis variabilis 2-oxoisovalerate + NH3 + H2O2
-
 ?

Subunits

EC Number Subunits Comment Organism
1.4.3.3 ? x * 38000, SDS-PAGE Trigonopsis variabilis

Synonyms

EC Number Synonyms Comment Organism
1.4.3.3 D-amino acid oxidase
-
 Trigonopsis variabilis
1.4.3.3 DAAO
-
 Trigonopsis variabilis

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.4.3.3 0.7
-
 D-Asn Km less than 0.7 s-1, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 1.75
-
 D-Thr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 3.54
-
 D-Lys wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 7.7
-
 D-Trp mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 11.5
-
 D-Val mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 13
-
 D-Asn mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 14.6
-
 D-Val mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 15.9
-
 D-Tyr mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 20.5
-
 D-Ser wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 22.5
-
 D-Tyr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 27.2
-
 D-Phe wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 29.1
-
 D-Leu wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 31
-
 D-Trp mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 33
-
 D-Trp mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 42
-
 D-Met mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 42.4
-
 D-Trp wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 46
-
 D-Leu mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 47.1
-
 D-Phe mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 50
-
 D-Asn mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 56
-
 D-Leu mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 58.8
-
 D-Phe mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 62.4
-
 D-Asn wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 63
-
 D-Met mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 63
-
 D-Phe mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 66.6
-
 D-Leu mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 72
-
 D-Tyr mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 80.5
-
 D-Met wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 85.3
-
 D-Val wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 89
-
 D-Met mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 108.6
-
 D-Ala wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis

Cofactor

EC Number Cofactor Comment Organism Structure
1.4.3.3 FAD
-
 Trigonopsis variabilis

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.4.3.3 0.12
-
 D-Lys wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.16
-
 D-Thr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.25
-
 D-Val mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.36
-
 D-Val mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.56
-
 D-Ser wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 0.59
-
 D-Asn mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 1.7
-
 D-Asn mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 2.8
-
 D-Asn wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 5.9
-
 D-Val wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 6.5
-
 D-Ala wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 12
-
 D-Trp mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 12.7
-
 D-Met mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 14.8
-
 D-Asn Km less than 14.8 mM-1 s-1, mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 17
-
 D-Trp mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 21
-
 D-Met mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 22
-
 D-Met mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 25
-
 D-Leu mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 37
-
 D-Phe mutant enzyme F258Y, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 37.3
-
 D-Leu wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 50
-
 D-Tyr wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 56
-
 D-Leu mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 67.3
-
 D-Trp mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 73.9
-
 D-Phe wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 85.8
-
 D-Tyr mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 86.5
-
 D-Trp wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 175
-
 D-Met wild type enzyme, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 183
-
 D-Tyr mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 245
-
 D-Leu mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 351
-
 D-Phe mutant enzyme F258S, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis
1.4.3.3 865
-
 D-Phe mutant enzyme F258A, in 50 mM potassium phosphate buffer, pH 8.0, at 30°C Trigonopsis variabilis