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Literature summary for 1.17.1.9 extracted from

  • Hoelsch, K.; Suehrer, I.; Heusel, M.; Weuster-Botz, D.
    Engineering of formate dehydrogenase: synergistic effect of mutations affecting cofactor specificity and chemical stability (2013), Appl. Microbiol. Biotechnol., 97, 2473-2481.
    View publication on PubMed
Search for more literature for 1.17.1.9

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Mycolicibacterium vaccae

Protein Variants

Protein Variants Comment Organism
A198G/D221Q the mutant has the highest catalytic efficiency (kcat/Km) with NADP+ Mycolicibacterium vaccae
C145S/A198G/D221Q/C255V the mutant confers a high resistance to ethyl 4-chloroacetoacetate (the half-life of this enzyme in 4 and 20 mM ethyl 4-chloroacetoacetate is over 200 h and 172 h, respectively) and shows 6fold enhanced catalytic efficiency with NADP+ compared to the wild type enzyme Mycolicibacterium vaccae
C145S/D221A/C255V the mutant has the lowest Km for formate Mycolicibacterium vaccae
C145S/D221Q/C255V the mutant shows the highest specific activity for a NADP+-accepting enzyme Mycolicibacterium vaccae
D221A the mutant shows catalytic efficiency with NADP+ as compared to the wild type enzyme Mycolicibacterium vaccae
D221G the mutant is extremely sensitive to ethyl 4-chloroacetoacetate with half-lives shorter than 6 min Mycolicibacterium vaccae
D221G/C255V the mutant is inhibited by NADP+ (Ki of 15 mM) Mycolicibacterium vaccae
D221Q the mutant shows catalytic efficiency with NADP+ compared to the wild type enzyme Mycolicibacterium vaccae
D221S the mutant shows catalytic efficiency with NADP+ as compared to the wild type enzyme Mycolicibacterium vaccae

Inhibitors

Inhibitors Comment Organism Structure
ethyl 4-chloroacetoacetate the D221G mutant is extremely sensitive to ethyl 4-chloroacetoacetate with half-lives shorter than 6 min Mycolicibacterium vaccae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.037
-
 NADP+ mutant enzyme D221A, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.04
-
 NADP+ Km above 0.04 mM, wild type enzyme, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.084
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.147
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
0.24
-
 NADP+ mutant enzyme D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.27
-
 NADP+ mutant enzyme A198G/D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.31
-
 NADP+ mutant enzyme D221G, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.39
-
 NADP+ mutant enzyme D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.43
-
 NADP+ mutant enzyme D221S, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.57
-
 NADP+ mutant enzyme C145S/D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.59
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.74
-
 NADP+ mutant enzyme C145S/D221A/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.92
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
1.09
-
 NAD+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
2 3 formate mutant enzyme D221A, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
4.1
-
 NAD+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
4.2
-
 formate wild type enzyme, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
40
-
 formate mutant enzyme D221G, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
41
-
 formate mutant enzyme D221S, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
46
-
 formate mutant enzyme D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
57
-
 formate mutant enzyme C145S/D221A/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
62
-
 formate mutant enzyme D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
69
-
 formate mutant enzyme C145S/D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
81
-
 formate mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
83
-
 formate mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
98
-
 formate mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
113
-
 formate mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
127
-
 formate mutant enzyme A198G/D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
CO2 + NADH Mycolicibacterium vaccae
-
 formate + NAD+
-
 ?
CO2 + NADH Mycolicibacterium vaccae N10
-
 formate + NAD+
-
 ?
formate + NAD+ Mycolicibacterium vaccae
-
 CO2 + NADH + H+
-
 ?
formate + NAD+ Mycolicibacterium vaccae N10
-
 CO2 + NADH + H+
-
 ?
formate + NADP+ Mycolicibacterium vaccae NADP+ is substrate for several mutant enzymes CO2 + NADPH
-
 ?
formate + NADP+ Mycolicibacterium vaccae N10 NADP+ is substrate for several mutant enzymes CO2 + NADPH
-
 ?

Organism

Organism UniProt Comment Textmining
Mycolicibacterium vaccae
-
-
-
Mycolicibacterium vaccae N10
-
-
-

Purification (Commentary)

Purification (Comment) Organism
HisTrap column chromatography Mycolicibacterium vaccae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
CO2 + NADH
-
 Mycolicibacterium vaccae formate + NAD+
-
 ?
CO2 + NADH
-
 Mycolicibacterium vaccae N10 formate + NAD+
-
 ?
formate + NAD+
-
 Mycolicibacterium vaccae CO2 + NADH + H+
-
 ?
formate + NAD+
-
 Mycolicibacterium vaccae N10 CO2 + NADH + H+
-
 ?
formate + NADP+ NADP+ is substrate for several mutant enzymes Mycolicibacterium vaccae CO2 + NADPH
-
 ?
formate + NADP+ NADP+ is substrate for several mutant enzymes Mycolicibacterium vaccae N10 CO2 + NADPH
-
 ?

Synonyms

Synonyms Comment Organism
FDH
-
 Mycolicibacterium vaccae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.52
-
 NADP+ mutant enzyme D221G, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.55
-
 NADP+ mutant enzyme D221S, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.62
-
 NADP+ mutant enzyme D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
0.79
-
 NADP+ mutant enzyme A198G/D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.04
-
 NADP+ mutant enzyme D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.05
-
 NADP+ mutant enzyme D221A, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.5
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.64
-
 NADP+ mutant enzyme C145S/D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.91
-
 NADP+ mutant enzyme C145S/D221A/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.76
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
3.08
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
5.18
-
 NAD+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
7.89
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
8.22
-
 NAD+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae

Cofactor

Cofactor Comment Organism Structure
NAD+
-
 Mycolicibacterium vaccae

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.29
-
 NADP+ mutant enzyme D221S, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
1.69
-
 NADP+ mutant enzyme D221G, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.58
-
 NADP+ mutant enzyme C145S/D221A/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.58
-
 NADP+ mutant enzyme D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.67
-
 NADP+ mutant enzyme D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.83
-
 NADP+ mutant enzyme D221A, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.88
-
 NADP+ mutant enzyme C145S/D221G/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
2.94
-
 NADP+ mutant enzyme A198G/D221Q, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
4.68
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
8.58
-
 NADP+ mutant enzyme C145S/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae
17.9
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 22°C Mycolicibacterium vaccae
21
-
 NADP+ mutant enzyme C145S/A198G/D221Q/C255V, in 0.1 M sodium phosphate buffer (pH 7.0), at 30°C Mycolicibacterium vaccae