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Literature summary extracted from

  • Abdallah, W.; Solanki, K.; Banta, S.
    Insertion of a calcium-responsive beta-roll domain into a thermostable alcohol dehydrogenase enables tunable control over cofactor selectivity (2018), ACS Catal., 8, 1602-1613 .
No PubMed abstract available

Protein Variants

EC Number Protein Variants Comment Organism
1.1.1.1 additional information insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity. Thus, the insertion of an intrinsically disordered calcium-binding domain into a key loop in a cofactor-dependent enzyme results in an enzyme with tunable cofactor selectivity, reminiscent of a calcium-controlled cofactor selectivity rheostat switch Pyrococcus furiosus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.1.1.1 additional information
-
 additional information kinetic data füor wild-type enzyme and chimeric enzyme created by insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D (AdhD) from Pyrococcus furiosus Pyrococcus furiosus

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.1.1.1 Ca2+ insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity Pyrococcus furiosus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
1.1.1.1 42000
-
 fusion protein, bet-AshD Pyrococcus furiosus

Organism

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

Purification (Commentary)

EC Number Purification (Comment) Organism
1.1.1.1
-
 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+ the enzyme is most active with 2,3-butanediol, preference for NADH in the reductive direction. Insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity Pyrococcus furiosus acetoin + NADH + H+
-
 ?
1.1.1.1 2,3-butanediol + NADP+ the enzyme is most active with 2,3-butanediol, preference for NADH in the reductive direction. Isertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity Pyrococcus furiosus acetoin + NADPH + H+
-
 ?

Subunits

EC Number Subunits Comment Organism
1.1.1.1 monomer 1 * 34000 Pyrococcus furiosus

Synonyms

EC Number Synonyms Comment Organism
1.1.1.1 AdhD
-
 Pyrococcus furiosus
1.1.1.1 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

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.1.1.1 additional information
-
 additional information kinetic data füor wild-type enzyme and chimeric enzyme created by insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D (AdhD) from Pyrococcus furiosus Pyrococcus furiosus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
1.1.1.1 9.3
-
 assay at Pyrococcus furiosus

Cofactor

EC Number Cofactor Comment Organism Structure
1.1.1.1 NAD+ insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity Pyrococcus furiosus
1.1.1.1 NADP+ insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D from Pyrococcus furiosus. The resultant chimera, beta-AdhD, gains the calcium-binding ability of the beta-roll, retains the thermostable activity of AdhD, and exhibits reduced overall alcohol dehydrogenase activity. The addition of calcium to beta-AdhD preferentially inhibits NAD+-dependent activity in comparison to NADP+-dependent activity. Calcium is a competitive inhibitor of AdhD, and the addition of the RTX domain introduces calcium-dependent noncompetitive inhibition to beta-AdhD affecting NAD+-dependent activity Pyrococcus furiosus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.1.1.1 additional information
-
 additional information kinetic data for wild-type enzyme and chimeric enzyme created by insertion of an RTX domain from the adenylate cyclase of Bordetella pertussis into a loop near the catalytic active site of the thermostable alcohol dehydrogenase D (AdhD) from Pyrococcus furiosus Pyrococcus furiosus