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

BRENDA support

Literature summary extracted from

  • Solanki, K.; Abdallah, W.; Banta, S.
    Engineering the cofactor specificity of an alcohol dehydrogenase via single mutations or insertions distal to the 2'-phosphate group of NADP(H) (2017), Protein Eng. Des. Sel., 30, 373-380 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.1.1.1 expression in Escherichia coli Pyrococcus furiosus

Protein Variants

EC Number Protein Variants Comment Organism
1.1.1.1 G211C mutant enzyme exhibits almost complete reversals in cofactor specificity Pyrococcus furiosus
1.1.1.1 G211InsA with NADP+, the mutant enzyme shows a 29-fold increase in kcat as compared to the wild-type enzyme Pyrococcus furiosus
1.1.1.1 G211InsC with NAD+, the mutant enzyme exhibits 6-fold decrease in kcat as compared to wild-type enzyme. Mutant enzyme exhibits almost complete reversals in cofactor specificity Pyrococcus furiosus
1.1.1.1 G211InsG with NAD+, the mutant enzyme exhibits 2.5-fold enhancement in kcat over the wild-type enzyme. Activity with NADP+ exceeds that of the wild type enzyme Pyrococcus furiosus
1.1.1.1 G211InsS with NAD+, the mutant enzyme exhibits 2.5-fold enhancement in kcat over the wild-type enzyme. With NADP+, the mutant enzyme shows a 29-fold increase in kcat as compared to the wild-type enzyme Pyrococcus furiosus
1.1.1.1 G211S with NAD+, the mutant enzyme exhibits 1.5-fold enhancement in kcat over the wild-type. Activity with NADP+ exceeds that of the wild type enzyme Pyrococcus furiosus
1.1.1.2 G211C site-directed mutagenesis, cofactor binding and kinetic analysis compared to the wild-type enzyme Pyrococcus furiosus
1.1.1.2 G211S site-directed mutagenesis, cofactor binding and kinetic analysis compared to the wild-type enzyme Pyrococcus furiosus
1.1.1.2 additional information insertion mutants of Ala, Gly, Ser or Cys between positions 211 and 212 (termed G211InsA, G211InsG, G211InsS, and G211InsC, respectively) are created by site-directed mutagenesis of the wild-type AdhD gene in the pET-20b vector. For some mutants (G211S, G211C, G211InsA and G211InsS), with NAD+ as a cofactor, the KA or KB values are observed to be higher than the maximum concentration of substrate or cofactor utilized. So, for mutants G211S, G211C, G211InsA and G211InsS, kinetic data are also measured using assay reaction mixtures containing 1-450 mM 2,3-butanediol and 1-5500 microM NAD+. In the case of NAD+, the on-rate of cofactor (kss 1 ) is found to decrease in all of the mutants except for the G211S and insertion of Gly (G211InsG). Kinetic analysis, detailed overview Pyrococcus furiosus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.1 0.097
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsC Pyrococcus furiosus
1.1.1.1 0.1
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211S Pyrococcus furiosus
1.1.1.1 0.12
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsA Pyrococcus furiosus
1.1.1.1 0.23
-
 NAD+ pH 8.8, 45°C, wild-type enzyme Pyrococcus furiosus
1.1.1.1 0.29
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsG Pyrococcus furiosus
1.1.1.1 0.44
-
 2,3-Butanediol pH 8.8, 45°C, wild-type enzyme, cofactor: NAP+ Pyrococcus furiosus
1.1.1.1 0.48
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211C Pyrococcus furiosus
1.1.1.1 0.91
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211S, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 1.5
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsG, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 1.6
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsS Pyrococcus furiosus
1.1.1.1 5.5
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsC, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 7.4
-
 NADP+ pH 8.8, 45°C, wild-type enzyme Pyrococcus furiosus
1.1.1.1 9.6
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211C, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 12
-
 2,3-Butanediol pH 8.8, 45°C, wild-type enzyme, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 15
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211S Pyrococcus furiosus
1.1.1.1 17
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsS, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 25
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211C Pyrococcus furiosus
1.1.1.1 33
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsG Pyrococcus furiosus
1.1.1.1 66
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsS Pyrococcus furiosus
1.1.1.1 76
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsC, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 90
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsA, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 100
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211S, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 110
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsG, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 120
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsC Pyrococcus furiosus
1.1.1.1 240
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211C, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 260
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsA, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 360
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsA Pyrococcus furiosus
1.1.1.1 1100
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsS, cofactor: NAD+ Pyrococcus furiosus
1.1.1.2 additional information
-
 additional information kinetic analysis of wild-type and mutant enzymes, fit to the ordered bi-bi rate equation, steady-state kinetics, detailed overview. For some mutants (G211S, G211C, G211InsA and G211InsS), with NAD+ as a cofactor, the KA or KB values are observed to be higher than the maximum concentration of substrate or cofactor utilized. So, for mutants G211S, G211C, G211InsA and G211InsS, kinetic data is also measured using assay reaction mixtures containing 1-450 mM 2,3-butanediol and 0.001-5.50 mM NAD+ Pyrococcus furiosus

Organism

EC Number Organism UniProt Comment Textmining
1.1.1.1 Pyrococcus furiosus
-
-
-
1.1.1.2 Pyrococcus furiosus
-
-
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.1.1.1 2,3-butanediol + NAD+
-
 Pyrococcus furiosus acetoin + NADH + H+
-
 ?
1.1.1.1 2,3-butanediol + NADP+
-
 Pyrococcus furiosus acetoin + NADPH + H+
-
 ?
1.1.1.2 2,3-butanediol + NADP+
-
 Pyrococcus furiosus acetoin + NADPH + H+
-
 r

Synonyms

EC Number Synonyms Comment Organism
1.1.1.1 AdhD
-
 Pyrococcus furiosus
1.1.1.1 alcohol dehydrogenase D
-
 Pyrococcus furiosus
1.1.1.2 AdhD
-
 Pyrococcus furiosus
1.1.1.2 alcohol dehydrogenase D
-
 Pyrococcus furiosus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
1.1.1.1 45
-
 assay at Pyrococcus furiosus
1.1.1.2 45
-
 assay at Pyrococcus furiosus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.1.1 0.073
-
 NADP+ pH 8.8, 45°C, wild-type enzyme Pyrococcus furiosus
1.1.1.1 0.073
-
 2,3-Butanediol pH 8.8, 45°C, wild-type enzyme, cofactor: NAP+ Pyrococcus furiosus
1.1.1.1 0.29
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211S Pyrococcus furiosus
1.1.1.1 0.29
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211S, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 0.43
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsC Pyrococcus furiosus
1.1.1.1 0.43
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsC, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 0.52
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsG Pyrococcus furiosus
1.1.1.1 0.52
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsG, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 0.86
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsC Pyrococcus furiosus
1.1.1.1 0.86
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsC, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 0.92
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211C Pyrococcus furiosus
1.1.1.1 0.92
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211C, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 1.1
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211C Pyrococcus furiosus
1.1.1.1 1.1
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211C, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 1.7
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsA Pyrococcus furiosus
1.1.1.1 1.7
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsA, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 2.1
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsA Pyrococcus furiosus
1.1.1.1 2.1
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsA, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 2.1
-
 NADP+ pH 8.8, 45°C, mutant enzyme G211InsS Pyrococcus furiosus
1.1.1.1 2.1
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsS, cofactor: NADP+ Pyrococcus furiosus
1.1.1.1 2.6
-
 NAD+ pH 8.8, 45°C, wild-type enzyme Pyrococcus furiosus
1.1.1.1 2.6
-
 2,3-Butanediol pH 8.8, 45°C, wild-type enzyme, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 4
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211S Pyrococcus furiosus
1.1.1.1 4
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211S, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 6.5
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsG Pyrococcus furiosus
1.1.1.1 6.5
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsG, cofactor: NAD+ Pyrococcus furiosus
1.1.1.1 6.7
-
 NAD+ pH 8.8, 45°C, mutant enzyme G211InsS Pyrococcus furiosus
1.1.1.1 6.7
-
 2,3-Butanediol pH 8.8, 45°C, mutant enzyme G211InsS, cofactor: NAD+ Pyrococcus furiosus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.1 8.8
-
 assay at Pyrococcus furiosus
1.1.1.2 8.8
-
 assay at Pyrococcus furiosus

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.1 NAD+
-
 Pyrococcus furiosus
1.1.1.1 NADP+
-
 Pyrococcus furiosus
1.1.1.2 additional information cofactor binding energies and kinetics with wild-type and mutant enzymes, overview Pyrococcus furiosus
1.1.1.2 NADP+
-
 Pyrococcus furiosus
1.1.1.2 NADPH
-
 Pyrococcus furiosus

General Information

EC Number General Information Comment Organism
1.1.1.2 additional information structure homology model of AdhD and its mutants generated using the model of prostaglandin F synthase from Trypanosoma brucei (PDB ID 1VBJ, 40.26% identity). The homology models are aligned with the structure of 2,5-DKGR from Corynebacterium (PDB ID 1A80 with bound NADPH, and PDB ID 1M9H with bound NADH), overview Pyrococcus furiosus