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Literature summary for 1.1.1.4 extracted from

  • Maddock, D.J.; Patrick, W.M.; Gerth, M.L.
    Substitutions at the cofactor phosphate-binding site of a clostridial alcohol dehydrogenase lead to unexpected changes in substrate specificity (2015), Protein Eng. Des. Sel., 28, 251-258.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli Clostridium autoethanogenum

Protein Variants

Protein Variants Comment Organism
G198D/S199 V/P201E/Y218A mutant displays no detectable activity with NADPH as cofactor, but uses NADH with kcat/Km value of 12 per s and mM Clostridium autoethanogenum
additional information variation of residue S199 does not yield any NADH-dependent enzymes Clostridium autoethanogenum
S199A increase in catalytic efficiencies Clostridium autoethanogenum
S199C decrease in catalytic efficiencies, with no significant changes in substrate preference Clostridium autoethanogenum
S199G decrease in catalytic efficiencies, with no significant changes in substrate preference Clostridium autoethanogenum
S199R increase in catalytic efficiencies Clostridium autoethanogenum
S199W decrease in catalytic efficiencies, with no significant changes in substrate preference Clostridium autoethanogenum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0017
-
NADPH wild-type, pH 7.5, 25°C Clostridium autoethanogenum
0.0056
-
NADPH mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
0.036
-
NADPH mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
0.8
-
2-butanone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
0.9
-
acetone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
1
-
acetone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
1.1
-
acetone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
1.2
-
acetone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
1.5
-
2-butanone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
1.6
-
acetone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
1.8
-
2-butanone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
1.9
-
2-butanone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
2
-
acetone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
2.4
-
2-butanone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
3.6
-
2-butanone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
34
-
(R)-acetoin mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
45
-
(R)-acetoin mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
98
-
(R)-acetoin wild-type, pH 7.5, 25°C Clostridium autoethanogenum
120
-
(R)-acetoin mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
190
-
(R)-acetoin mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
230
-
(R)-acetoin mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum

Organism

Organism UniProt Comment Textmining
Clostridium autoethanogenum F8TEL7
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(R)-acetoin + NADPH + H+
-
Clostridium autoethanogenum (2R,3R)-2,3-butanediol + NAD+
-
?
2-butanone + NADP+
-
Clostridium autoethanogenum 2-butanol + NADPH + H+
-
?
acetone + NADPH + H+
-
Clostridium autoethanogenum propan-2-ol + NADP+
-
?

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2 8 2-butanone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
3 6 acetone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
23
-
2-butanone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
32
-
NADPH mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
37
-
2-butanone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
40
-
2-butanone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
41
-
(R)-acetoin mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
46
-
2-butanone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
48
-
acetone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
50
-
NADPH mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
50
-
2-butanone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
55
-
acetone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
60
-
acetone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
62
-
acetone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
66
-
(R)-acetoin mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
68
-
NADPH wild-type, pH 7.5, 25°C Clostridium autoethanogenum
89
-
acetone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
95
-
(R)-acetoin mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
110
-
(R)-acetoin mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
150
-
(R)-acetoin wild-type, pH 7.5, 25°C Clostridium autoethanogenum
230
-
(R)-acetoin mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum

Cofactor

Cofactor Comment Organism Structure
NADH cofoactor of mutant G198D/S199 V/P201E/Y218A Clostridium autoethanogenum
NADPH strictly dependent on NADPH Clostridium autoethanogenum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.35
-
(R)-acetoin mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
0.48
-
(R)-acetoin mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
0.9
-
(R)-acetoin mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
1.6
-
(R)-acetoin wild-type, pH 7.5, 25°C Clostridium autoethanogenum
1.9
-
(R)-acetoin mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
2.8
-
(R)-acetoin mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
7.8
-
2-butanone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
15
-
2-butanone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
17
-
2-butanone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
21
-
2-butanone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
24
-
acetone mutant S199W, pH 7.5, 25°C Clostridium autoethanogenum
26
-
2-butanone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
30
-
acetone mutant S199C, pH 7.5, 25°C Clostridium autoethanogenum
39
-
acetone mutant S199G, pH 7.5, 25°C Clostridium autoethanogenum
55
-
acetone wild-type, pH 7.5, 25°C Clostridium autoethanogenum
61
-
2-butanone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
67
-
acetone mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
81
-
acetone mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
890
-
NADPH mutant S199R, pH 7.5, 25°C Clostridium autoethanogenum
8900
-
NADPH mutant S199A, pH 7.5, 25°C Clostridium autoethanogenum
39000
-
NADPH wild-type, pH 7.5, 25°C Clostridium autoethanogenum