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Literature summary for 1.1.1.184 extracted from

  • Zhang, R.; Zhu, G.; Zhang, W.; Cao, S.; Ou, X.; Li, X.; Bartlam, M.; Xu, Y.; Zhang, X.C.; Rao, Z.
    Crystal structure of a carbonyl reductase from Candida parapsilosis with anti-Prelog stereospecificity (2008), Protein Sci., 17, 1412-1423.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
SCR overexpressed from pETSCR plasmid in Escherichia coli BL21 (DE3) as His6-tagged proteins Candida parapsilosis

Crystallization (Commentary)

Crystallization (Comment) Organism
by the hanging-drop vapor diffusion method, to 2.7 A resolution, crystal form belongs to space group P212121 with cell dimensions of a=104.7 A, b=142.8 A, and c=151.8 A. It forms a homotetramer with a broken 2-2-2 symmetry. SCR contains an extended N-terminal peptide (i.e., residues 1-25) and a short helix (residues 26-30) projecting out from the core domain that may stabilize the oligomer. In the apo-SCR structure, the entrance of the NADPH pocket is blocked by a surface loop Candida parapsilosis

Protein Variants

Protein Variants Comment Organism
DELTA31 deletion of the N-terminal 31 residues, kinetic parameters KM and kcat are essentially the same as those of the wild-type. It forms a tetramer in solution, which is similar to the wild-type but is less stable. Melting temperature is 48°C, which is lower than that of the wild-type (52°C) Candida parapsilosis
S172A melting temperature is 46°C, which is lower than that of the wild-type (52°C). Shows no activity Candida parapsilosis
S172T melting temperature is 48°C, which is lower than that of the wild-type (52°C). Activity is essentially the same as the wild-type Candida parapsilosis
S67D/H68D double-point mutation inside the coenzyme-binding pocket results in a nearly 10fold increase and a 20fold decrease in the kcat/KM value when NADH and NADPH are used as cofactors, respectively, with kcat remaining essentially the same. Shows similar thermal stability to wild-type Candida parapsilosis
V270D renders the SCR as a homodimer, rather than a tetramer, without affecting the enzymatic activity. Melting temperature is 45°C, which is lower than that of the wild-type (52°C) Candida parapsilosis
Y187A melting temperature is 45°C, which is lower than that of the wild-type (52°C). Shows no activity Candida parapsilosis
Y187F melting temperature is 46°C, which is lower than that of the wild-type (52°C). Shows no activity Candida parapsilosis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.59
-
2-hydroxyacetophenone wild-type, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
0.63
-
2-hydroxyacetophenone mutant DELTA31, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
0.68
-
2-hydroxyacetophenone mutant S172T, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
0.71
-
2-hydroxyacetophenone mutant V270D, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
0.76
-
2-hydroxyacetophenone mutant S67D/H68D, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
7.89
-
2-hydroxyacetophenone wild-type, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
8.03
-
2-hydroxyacetophenone mutant DELTA31, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
8.45
-
2-hydroxyacetophenone mutant V270D, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
8.67
-
2-hydroxyacetophenone mutant S172T, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
10.27
-
2-hydroxyacetophenone mutant S67D/H68D, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
58000
-
gel filtration, mutant V270D Candida parapsilosis
100000
-
analytic ultracentrifugation, mutant DELTA31 Candida parapsilosis
112000
-
analytic ultracentrifugation, wild-type Candida parapsilosis

Organism

Organism UniProt Comment Textmining
Candida parapsilosis B2KJ46
-
-

Purification (Commentary)

Purification (Comment) Organism
by affinity chromatography on a Ni2+ column and by gel filtration Candida parapsilosis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2-hydroxyacetophenone + NADPH + H+
-
Candida parapsilosis (S)-1-phenyl-1,2-ethanediol + NADP+
-
?

Subunits

Subunits Comment Organism
homodimer analytic ultracentrifugation analysis, mutant V270D Candida parapsilosis
homotetramer crystallography, wild-type Candida parapsilosis

Synonyms

Synonyms Comment Organism
SCR
-
Candida parapsilosis
short-chain (S)-1-phenyl-1,2-ethanediol dehydrogenase
-
Candida parapsilosis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.13
-
2-hydroxyacetophenone mutant S172T, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.21
-
2-hydroxyacetophenone mutant S67D/H68D, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.22
-
2-hydroxyacetophenone mutant V270D, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.24
-
2-hydroxyacetophenone mutant DELTA31, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.26
-
2-hydroxyacetophenone mutant S67D/H68D, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.32
-
2-hydroxyacetophenone wild-type, with NADH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.34
-
2-hydroxyacetophenone mutant V270D, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.36
-
2-hydroxyacetophenone mutant S172T, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.39
-
2-hydroxyacetophenone mutant DELTA31, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis
1.48
-
2-hydroxyacetophenone wild-type, with NADPH as cofactor, at 35°C in 100 mM acetate buffer (pH 5.0) Candida parapsilosis

Cofactor

Cofactor Comment Organism Structure
NAD(P)H the wild-type exhibits coenzyme specificity for NADPH over NADH, whereas NADH has stronger inducing ability than NADPH for the S67D/H68D mutant Candida parapsilosis