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Literature summary for 1.1.1.2 extracted from

  • Xu, J.; Zhou, H.; Yu, H.; Deng, T.; Wang, Z.; Zhang, H.; Wu, J.; Yang, L.
    Computational design of highly stable and soluble alcohol dehydrogenase for NADPH regeneration (2021), Bioresour. Bioprocess., 8, 12 .
No PubMed abstract available
Search for more literature for 1.1.1.2

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3) Clostridium beijerinckii

Protein Variants

Protein Variants Comment Organism
additional information computational design of highly stable and soluble alcohol dehydrogenase mutants for NADPH regeneration, several mutants are created and evaluated, overview. A NADP+-specific ADH from Clostridium beijerinckii is engineered for cofactor recycling using an automated algorithm. The mutant is selected for large-scale production and industrial usage Clostridium beijerinckii
S24P/G182A/G196A/H222D/S250E/S254R the mutant CbADH-6M exhibits a favorable soluble and highly active expression with an activity of 46.3 U/ml, which is 16times higher than the wild-type (2.9 U/ml) and a more stable protein conformation with an enhanced thermal stability. The activity of CbADH-6M is upgraded to 2401.8 U/ml by high cell density fermentation strategy using recombinant Escherichia coli. Improving CbADH solubility by chaperone buffering. The efficiency for NADPH regeneration of the mutant enzyme is testified in the synthesis of some fine chiral aromatic alcohols coupling with another ADH from Lactobacillus kefir (LkADH). Method optimization, overview. Although the Asp225-His222 salt bridge in the original wild-type CbADH protein disappears due to the amino acid substitution H222D, the replacement of H222D, S250E and S254R leads to the formation of four new salt bridges including Arg254-Glu250, Arg254-Asp225, Arg254-Glu280 and Arg254-Asp222, which constitute a salt bridge network centered on Arg254 Clostridium beijerinckii

Localization

Localization Comment Organism GeneOntology No. Textmining
soluble
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 Clostridium beijerinckii
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Metals/Ions

Metals/Ions Comment Organism Structure
Zn2+ required Clostridium beijerinckii

Organism

Organism UniProt Comment Textmining
Clostridium beijerinckii
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-
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Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
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 activity of the crude recombinant mutant CbADH-6M is 46.3 U/ml, that of the crude recombinant wild-type enzyme is 2.9 U/ml Clostridium beijerinckii

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1-(3,4,5-trifluorophenyl)ethan-1-one + NADPH + H+
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 Clostridium beijerinckii (1S)-1-(3,4,5-trifluorophenyl)ethan-1-ol + NADP+ 90.8% conversion, 99% ee r
1-(3-chlorophenyl)ethan-1-one + NADPH + H+
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 Clostridium beijerinckii (1S)-1-(3-chlorophenyl)ethan-1-ol + NADP+ 99.5% conversion, 99% ee r
1-(4-fluorophenyl)ethan-1-one + NADPH + H+
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 Clostridium beijerinckii (1S)-1-(4-fluorophenyl)ethan-1-ol + NADP+ 99.5% conversion, 99% ee r
1-(4-methylphenyl)ethan-1-one + NADPH + H+
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 Clostridium beijerinckii (1S)-1-(4-methylphenyl)ethan-1-ol + NADP+ 99.5% conversion, 99% ee r
1-phenylethan-1-one + NADPH + H+
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 Clostridium beijerinckii (1S)-1-phenylethan-1-ol + NADP+ 99.5% conversion, 99% ee r
additional information the enatiomeric excess with all substrates is over 99% Clostridium beijerinckii ?
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Subunits

Subunits Comment Organism
dimer
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 Clostridium beijerinckii

Synonyms

Synonyms Comment Organism
CbADH
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 Clostridium beijerinckii
NADPH-dependent ADH
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 Clostridium beijerinckii

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
35
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 assay at Clostridium beijerinckii

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
63.7
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 half-life of the wild-type enzyme is 60 min Clostridium beijerinckii
66.8
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 half-life of the enzyme mutant CbADH-6M is 60 min Clostridium beijerinckii

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
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 assay at Clostridium beijerinckii

Cofactor

Cofactor Comment Organism Structure
NADP+
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 Clostridium beijerinckii
NADPH
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 Clostridium beijerinckii