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

BRENDA support

Literature summary for 1.1.1.B4 extracted from

  • Protsko, C.; Vieille, C.; Laivenieks, M.; Prasad, L.; Sanders, D.A.; Delbaere, L.T.
    Crystallization and preliminary X-ray diffraction analysis of the Thermoanaerobacter ethanolicus secondary alcohol dehydrogenase I86A mutant (2010), Acta Crystallogr. Sect. F, 66, 831-833.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene adhB, expression of mutant I86A in Escherichia coli HB101 (DE3) as protein C-terminally tagged with two methionines followed by Leu-Glu-His6 Thermoanaerobacter ethanolicus

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant His-tagged mutant I86A, hanging drop vapour diffusion, mixing of 0.002 ml of 8 mg/ml protein in 50 mM 2-(N-morpholino)ethanesulfonic acid, pH 6.6, 20 mM NaCl, 0.1 mM dithiothreitol, 0.1 mM ethylenediaminetetraacetic acid, and 2 mM (R)-1-phenylethanol, with 0.2% D-fructose 1,6-disphosphate trisodium salt octahydrate, 0.2% glycerol phosphate disodium salt hydrate, 0.2% O-phospho-L-serine, 0.2% O-phosphol-L-styrosine, 0.2% phytic acid sodium salt hydrate, 0.02 M HEPES sodium pH 6.8, and 0.002 ml reservoir solution containing 20% v/v PEG 400, 0.1 M Na HEPES, pH 7.0, room temperature, 6-12 h, X-ray diffraction structure determination and analysis, molecular replacement, modeling Thermoanaerobacter ethanolicus

Protein Variants

Protein Variants Comment Organism
I86A site-directed mutagensis, the secondary alcohol dehydrogenase I86A mutant is stereospecific for (R)-alcohols instead of (S)-alcohols, in contrast to the wild-type enzyme, the mutation I86A allows large substituents to fit into the large pocket of I86ATeSADH, which corresponds to the small pocket in wild-type TeSADH, modeling of the stereopreference of TeSADH I86A Thermoanaerobacter ethanolicus

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
37590
-
4 * 37590, sequence calculation, a tetramer formed by a dimer of dimers Thermoanaerobacter ethanolicus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
(R)-1-phenylethanol + NAD+ Thermoanaerobacter ethanolicus activity of mutant I886A, the wild-type enzyme is (S)-specific forming (S)-1-phenylethanol acetophenone + NADH + H+
-
r

Organism

Organism UniProt Comment Textmining
Thermoanaerobacter ethanolicus P77990 gene adhB, mutant I86A
-

Purification (Commentary)

Purification (Comment) Organism
recombinant C-terminally His-tagged mutant I86A from Escherichia coli HB101 (DE3) by nickel affinity chromatography Thermoanaerobacter ethanolicus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(R)-1-phenylethanol + NAD+ activity of mutant I886A, the wild-type enzyme is (S)-specific forming (S)-1-phenylethanol Thermoanaerobacter ethanolicus acetophenone + NADH + H+
-
r

Subunits

Subunits Comment Organism
tetramer 4 * 37590, sequence calculation, a tetramer formed by a dimer of dimers Thermoanaerobacter ethanolicus

Cofactor

Cofactor Comment Organism Structure
NAD+
-
Thermoanaerobacter ethanolicus
NADH
-
Thermoanaerobacter ethanolicus

General Information

General Information Comment Organism
additional information the mutation I86A allows large substituents to fit into the large pocket of I86ATeSADH, which corresponds to the small pocket in wild-type TeSADH, modeling of the stereopreference of TeSADH I86A Thermoanaerobacter ethanolicus