Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Xu, Y.; Dewanti, A.R.; Mitra, B.
    Arginine 165/arginine 277 pair in (S)-mandelate dehydrogenase from Pseudomonas putida: role in catalysis and substrate binding (2002), Biochemistry, 41, 12313-12319.
    View publication on PubMed

Protein Variants

EC Number Protein Variants Comment Organism
1.1.99.31 R165E mutant is less stable than wild-type enzyme, mutant enzyme is only partially reduced by (S)-mandelate. 81.8fold decrease in kcat for (S)-mandelate, 199fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R165G mutant is less stable than wild-type enzyme, mutant enzyme is only partially reduced by (S)-mandelate. 27.3fold decrease in kcat for (S)-mandelate, 48.3fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R165G/R277G no activity. Double mutant is less stable than single Arg165 mutant in term of long-term storage Pseudomonas putida
1.1.99.31 R165G/R277K double mutant is less stable than single Arg165 mutant in term of long-term storage. 9000fold decrease in kcat for (S)-mandelate, 261.7fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R165K mutant is less stable than wild-type enzyme, mutant enzyme is fully reduced on addition of (S)-mandelate. 3fold decrease in kcat for (S)-mandelate, 12.5fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R165K/R277G no activity. Double mutant is less stable than single Arg165 mutant in term of long-term storage Pseudomonas putida
1.1.99.31 R165K/R277K double mutant is less stable than single Arg165 mutant in term of long-term storage. 327fold decrease in kcat for (S)-mandelate, 126.7fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R165M mutant is less stable than wild-type enzyme, mutant enzyme is fully reduced on addition of (S)-mandelate. 19.6fold decrease in kcat for (S)-mandelate, 36.7fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R277G 1565fold decrease in kcat for (S)-mandelate, 141fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida
1.1.99.31 R277K 5.5 fold decrease in kcat for (S)-mandelate, 46.7fold increase in KM-value for (S)-mandelate compared to wild-type enzyme Pseudomonas putida

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.1.99.31 (R)-mandelate
-
 Pseudomonas putida
1.1.99.31 2-phenylethanoate
-
 Pseudomonas putida
1.1.99.31 phenylethanediol poor competitive inhibitor of wild-type enzyme Pseudomonas putida

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.99.31 0.12
-
 (S)-Mandelate pH 7.5, 20°C, wild-type enzyme Pseudomonas putida
1.1.99.31 1.5
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165K Pseudomonas putida
1.1.99.31 4.4
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165M Pseudomonas putida
1.1.99.31 5.6
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R277K Pseudomonas putida
1.1.99.31 5.8
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165G Pseudomonas putida
1.1.99.31 12
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165E Pseudomonas putida
1.1.99.31 15.2
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165K/R277K Pseudomonas putida
1.1.99.31 17
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R277G Pseudomonas putida
1.1.99.31 31.4
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165G/R277K Pseudomonas putida

Organism

EC Number Organism UniProt Comment Textmining
1.1.99.31 Pseudomonas putida
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.99.31
-
 Pseudomonas putida

Storage Stability

EC Number Storage Stability Organism
1.1.99.31 -70°C, wild-type enzyme retains activity for more than 1 year. The single Arg165 mutant lose activity after 2-3 months, double mutants R165K/R277K and R165G/R277K are inactivated after 2-3 weeks in storage Pseudomonas putida

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.99.31 (S)-mandelate + acceptor reduction of 2,6-dichloroindophenol in presence of N-methylphenazonium methosulfate Pseudomonas putida 2-oxo-2-phenylacetate + reduced acceptor
-
 ?

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.99.31 0.04
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165G/R277K Pseudomonas putida
1.1.99.31 0.23
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R277G Pseudomonas putida
1.1.99.31 1.1
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165K/R277K Pseudomonas putida
1.1.99.31 4.4
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165E Pseudomonas putida
1.1.99.31 13.2
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165G Pseudomonas putida
1.1.99.31 18.4
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165M Pseudomonas putida
1.1.99.31 66
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R277K Pseudomonas putida
1.1.99.31 120
-
 (S)-Mandelate pH 7.5, 20°C, mutant enzyme R165K Pseudomonas putida
1.1.99.31 360
-
 (S)-Mandelate pH 7.5, 20°C, wild-type enzyme Pseudomonas putida

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.99.31 FMN
-
 Pseudomonas putida

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.1.99.31 2.9
-
 (R)-mandelate pH 7.5, 20°C, wild-type enzyme Pseudomonas putida
1.1.99.31 3.1
-
 (R)-mandelate pH 7.5, 20°C, mutant enzyme R165K Pseudomonas putida
1.1.99.31 7.2
-
 (R)-mandelate pH 7.5, 20°C, mutant enzyme R277K Pseudomonas putida
1.1.99.31 11
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R165K Pseudomonas putida
1.1.99.31 11
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R165K/R277K Pseudomonas putida
1.1.99.31 12
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R277K Pseudomonas putida
1.1.99.31 12
-
 2-phenylethanoate pH 7.5, 20°C, wild-type enzyme Pseudomonas putida
1.1.99.31 19.5
-
 (R)-mandelate pH 7.5, 20°C, mutant enzyme R277G Pseudomonas putida
1.1.99.31 20.3
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R165G/R277K Pseudomonas putida
1.1.99.31 22.5
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R277G Pseudomonas putida
1.1.99.31 67
-
 2-phenylethanoate pH 7.5, 20°C, mutant enzyme R165G Pseudomonas putida
1.1.99.31 73
-
 (R)-mandelate pH 7.5, 20°C, mutant enzyme R165G Pseudomonas putida
1.1.99.31 700
-
 (R)-mandelate pH 7.5, 20°C, mutant enzyme R165G/R277K Pseudomonas putida