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Literature summary for 1.1.1.50 extracted from

  • Hwang, C.; Chang, Y.; Hsu, C.; Hsu, H.; Li, C.; Pon, H.
    Mechanistic roles of Ser-114, Tyr-155, and Lys-159 in 3alpha-hydroxysteroid dehydrogenase/carbonyl reductase from Comamonas testosteroni (2005), J. Biol. Chem., 280, 3522-3528.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
overexpression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Comamonas testosteroni

Protein Variants

Protein Variants Comment Organism
K159A site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 200fold reduced activity compared to the wild-type Comamonas testosteroni
S114A site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 3400fold reduced activity compared to the wild-type Comamonas testosteroni
S114A/Y155F site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 200000fold reduced activity compared to the wild-type Comamonas testosteroni
Y155F site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 2800fold reduced activity compared to the wild-type Comamonas testosteroni
Y155F/K159A site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 9400fold reduced activity compared to the wild-type Comamonas testosteroni

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information steady-state kinetics Comamonas testosteroni
0.00044
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant S114A/Y155F Comamonas testosteroni
0.0055
-
androsterone pH 9.0, 25°C, recombinant His-tagged wild-type Comamonas testosteroni
0.007
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant S114A Comamonas testosteroni
0.065
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant K159A Comamonas testosteroni
0.074
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant Y155F Comamonas testosteroni
0.1
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant Y155F/K159A Comamonas testosteroni

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
70000
-
about, recombinant His-tagged wild-type and mutant enzymes, gel filtration Comamonas testosteroni

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
androsterone + NAD+ Comamonas testosteroni
-
5alpha-androstane-3,17-dione + NADH
-
r

Organism

Organism UniProt Comment Textmining
Comamonas testosteroni
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, and gel filtration to homogeneity Comamonas testosteroni

Reaction

Reaction Comment Organism Reaction ID
a 3alpha-hydroxysteroid + NAD(P)+ = a 3-oxosteroid + NAD(P)H + H+ the catalytic triad is composed of Ser-114, Tyr-155, and Lys-159, structural analysis and sequence alignment of the short chain dehydrogenase/reductase family Comamonas testosteroni

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
-
Comamonas testosteroni

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
androsterone + NAD+
-
Comamonas testosteroni 5alpha-androstane-3,17-dione + NADH
-
r
additional information bifunctional enzyme also catalyzing the reaction of carbonyl reductase, EC 1.1.1.184 Comamonas testosteroni ?
-
?

Subunits

Subunits Comment Organism
dimer 2 * 28349-28425, recombinant His-tagged wild-type and mutant enzymes, mass spectrometry Comamonas testosteroni

Synonyms

Synonyms Comment Organism
3alpha-HSD/CR
-
Comamonas testosteroni
3alpha-hydroxysteroid dehydrogenase/carbonyl reductase
-
Comamonas testosteroni
More cf. EC 1.1.1.184 Comamonas testosteroni

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Comamonas testosteroni

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0028
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant S114A/Y155F Comamonas testosteroni
0.003
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant Y155F/K159A Comamonas testosteroni
0.12
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant S114A Comamonas testosteroni
1.5
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant Y155F Comamonas testosteroni
18.9
-
androsterone pH 9.0, 25°C, recombinant His-tagged mutant K159A Comamonas testosteroni
318
-
androsterone pH 9.0, 25°C, recombinant His-tagged wild-type Comamonas testosteroni

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
9
-
assay at Comamonas testosteroni

pH Range

pH Minimum pH Maximum Comment Organism
5 11 pH profile, recombinant His-tagged wild-type and mutant enzymes Comamonas testosteroni

Cofactor

Cofactor Comment Organism Structure
NAD+
-
Comamonas testosteroni
NADH
-
Comamonas testosteroni