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Literature summary for 1.3.1.85 extracted from

  • Stoffel, G.M.M.; Saez, D.A.; DeMirci, H.; Voegeli, B.; Rao, Y.; Zarzycki, J.; Yoshikuni, Y.; Wakatsuki, S.; Voehringer-Martinez, E.; Erb, T.J.
    Four amino acids define the CO2 binding pocket of enoyl-CoA carboxylases/reductases (2019), Proc. Natl. Acad. Sci. USA, 116, 13964-13969 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.3.1.85

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21 AI (DE3) cells Kitasatospora setae

Crystallization (Commentary)

Crystallization (Comment) Organism
apoenzyme and in complex with NADPH and (2S)-ethylmalonyl-CoA, sitting drop vapor diffusion method, using 100 mM Tris pH 8.0 and 20% (w/v) poly (acrylic acid sodium salt) 5100 Kitasatospora setae

Protein Variants

Protein Variants Comment Organism
E171A the mutant shows reduced activity compared to the wild type enzyme Kitasatospora setae
F170A in the mutant, the carboxylation activity is decreased to 17% at the expense of increased reduction side reactivity compared to the wild type enzyme Kitasatospora setae
F170Y the mutant shows a slightly increased substrate inhibition, but otherwise very similar kinetic parameters as the wild type enzyme Kitasatospora setae
H365N the mutant shows a 20fold decreased activity compared with the wild type enzyme but still displays 93% of carboxylated product, even though the KM for CO2 is raised by more than 1 order of magnitude Kitasatospora setae
N81L the interaction of the enzyme with CO2 is severely disturbed by the N81L mutation. Stereospecificity of butyryl-CoA formation is almost completely lost in the mutant compared with the wild type enzyme Kitasatospora setae
S119A the mutant shows almost full carboxylation, but at more than 50fold reduced turnover frequency as compared to the wild type enzyme Kitasatospora setae
T82D the mutant shows almost full carboxylation, but at more than 50fold reduced turnover frequency as compared to the wild type enzyme Kitasatospora setae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.01
-
 crotonyl-CoA mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
0.011
-
 NADPH mutant enzyme F170A, at pH 8.0 and 30°C Kitasatospora setae
0.0134
-
 crotonyl-CoA mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
0.021
-
 crotonyl-CoA wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae
0.022
-
 NADPH mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae
0.0298
-
 crotonyl-CoA mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae
0.03
-
 NADPH mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
0.03
-
 CO2 mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
0.031
-
 crotonyl-CoA mutant enzyme F170A, at pH 8.0 and 30°C Kitasatospora setae
0.036
-
 NADPH mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
0.036
-
 crotonyl-CoA mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
0.037
-
 NADPH wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae
0.052
-
 NADPH mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
0.08
-
 CO2 mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
0.09
-
 CO2 wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae
0.112
-
 NADPH mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
0.15
-
 CO2 mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
0.155
-
 CO2 mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
0.5
-
 crotonyl-CoA mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
1.31
-
 CO2 mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae

Organism

Organism UniProt Comment Textmining
Kitasatospora setae E4N096
-
-
Kitasatospora setae ATCC 33774 E4N096
-
-

Purification (Commentary)

Purification (Comment) Organism
HisTrap column chromatography Kitasatospora setae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
crotonyl-CoA + CO2 + NADPH + H+
-
 Kitasatospora setae (2S)-ethylmalonyl-CoA + NADP+
-
 ?
crotonyl-CoA + CO2 + NADPH + H+
-
 Kitasatospora setae ATCC 33774 (2S)-ethylmalonyl-CoA + NADP+
-
 ?
additional information the enzyme catalyzes the reduction of crotonyl-CoA to butyryl-CoA as a side reaction, but only in the absence of CO2 and at low catalytic efficiency Kitasatospora setae ?
-
-
additional information the enzyme catalyzes the reduction of crotonyl-CoA to butyryl-CoA as a side reaction, but only in the absence of CO2 and at low catalytic efficiency Kitasatospora setae ATCC 33774 ?
-
-

Synonyms

Synonyms Comment Organism
CCR
-
 Kitasatospora setae
ECR
-
 Kitasatospora setae
enoyl-CoA carboxylase/reductase
-
 Kitasatospora setae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.157
-
 NADPH mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
0.17
-
 CO2 mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
0.172
-
 crotonyl-CoA mutant enzyme T82D, at pH 8.0 and 30°C Kitasatospora setae
1.32
-
 CO2 mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
1.47
-
 NADPH mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
1.79
-
 crotonyl-CoA mutant enzyme S119A, at pH 8.0 and 30°C Kitasatospora setae
5
-
 crotonyl-CoA mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae
5.1
-
 CO2 mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
5.1
-
 crotonyl-CoA mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
6
-
 NADPH mutant enzyme E171A, at pH 8.0 and 30°C Kitasatospora setae
7.4
-
 CO2 mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae
8.1
-
 NADPH mutant enzyme H365N, at pH 8.0 and 30°C Kitasatospora setae
8.3
-
 crotonyl-CoA mutant enzyme F170A, at pH 8.0 and 30°C Kitasatospora setae
11
-
 NADPH mutant enzyme F170A, at pH 8.0 and 30°C Kitasatospora setae
56
-
 NADPH mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
56
-
 CO2 mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
78
-
 CO2 wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae
83
-
 crotonyl-CoA mutant enzyme F170Y, at pH 8.0 and 30°C Kitasatospora setae
86
-
 NADPH wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae
103
-
 crotonyl-CoA wild type enzyme, at pH 8.0 and 30°C Kitasatospora setae

Cofactor

Cofactor Comment Organism Structure
NADPH
-
 Kitasatospora setae