Short-chain alcohol dehydrogenase from Thermus thermophilus HB27

Enzyme Description

Species

Thermus thermophilus HB27 ↗

Extremophile

Yes "extremely thermophilic" organism

EC Number

1.1.1.- ↗

Sequence

Length: 256 amino acids

MGLFAGKGVLVTGGARGIGRAIAQAFAREGALVALCDLRPEGKEVAEAIGGAFFQVDLEDERERVRFVEEAAYALGRVDVLVNNAAIAAPGSALTVRLPEWRRVLEVNLTAPMHLSALAAREMRKVGGGAIVNVASVQGLFAEQENAAYNASKGGLVNLTRSLALDLAPLRIRVNAVAPGAIATEAVLEAIALSPDPERTRRDWEDLHALRRLGKPEEVAEAVLFLASEKASFITGAILPVDGGMTASFMMAGRPV

Angela Pennacchio et al. (2008) — Purification and Characterization of a Novel Recombinant Highly Enantioselective Short-Chain NAD(H)-Dependent Alcohol Dehydrogenase from Thermus thermophilus

Applied and Environmental Microbiology · doi:10.1128/AEM.00217-08 ↗ · Activity + Stability - Incubation

24 measurements ▼

Database ID

UniProt: A0A3P4ASV8 ↗

Sequence Annotation

Explicit - Provided

Protein Source

Recombinant, host bacterium Escherichia coli BL21 (DE3)

Experimental Data (24 measurements)

24 measurements — page 2 of 2

Property	Assay	Solvent	Solvent Volume	Aqueous Reference	Measured Value	Units	Solution	pH	Temperature	Substrate(s)	Product(s)	Cofactor(s)	Shaking	Comments
Activity + Stability - Incubation	Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent	Isopropanol	2%	130	165	%	Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl	Incubation: 6, Assay: 6	Incubation: 50°C, Assay: 65°C	20 mM Ethyl benzoylformate	Ethyl (R)-mandelate (95% ee)	0.3 mM NADH	—	Water-incubated control (in %), activity measured in the presence of organic solvent \| Not completely sure if the assay in the presence of organic solvent ("The concentration of the additive during the standard assay was as follows: 10 mM inhibitor, 10% detergent, and 50% (v/v) or 90% (v/v) organic solvent.")
Activity + Stability - Incubation	Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent	Methanol	5%	134	186	%	Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl	Incubation: 6, Assay: 6	Incubation: 60°C, Assay: 65°C	20 mM Ethyl benzoylformate	Ethyl (R)-mandelate (95% ee)	0.3 mM NADH	—	Water-incubated control (in %), activity measured in the presence of organic solvent \| Not completely sure if the assay in the presence of organic solvent ("The concentration of the additive during the standard assay was as follows: 10 mM inhibitor, 10% detergent, and 50% (v/v) or 90% (v/v) organic solvent.")
Activity + Stability - Incubation	Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent	Methanol	5%	129	186	%	Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl	Incubation: 6, Assay: 6	Incubation: 60°C, Assay: 65°C	20 mM Ethyl benzoylformate	Ethyl (R)-mandelate (95% ee)	0.3 mM NADH	—	Water-incubated control (in %), activity measured in the presence of organic solvent \| Not completely sure if the assay in the presence of organic solvent ("The concentration of the additive during the standard assay was as follows: 10 mM inhibitor, 10% detergent, and 50% (v/v) or 90% (v/v) organic solvent.")
Activity + Stability - Incubation	Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent	Methanol	5%	142	137	%	Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl	Incubation: 6, Assay: 6	Incubation: 50°C, Assay: 65°C	20 mM Ethyl benzoylformate	Ethyl (R)-mandelate (95% ee)	0.3 mM NADH	—	Water-incubated control (in %), activity measured in the presence of organic solvent \| Not completely sure if the assay in the presence of organic solvent ("The concentration of the additive during the standard assay was as follows: 10 mM inhibitor, 10% detergent, and 50% (v/v) or 90% (v/v) organic solvent.")

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Acetonitrile

130

131

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Water-incubated control (in %), activity measured in the presence of organic solvent | Not completely sure if the assay in the presence of organic solvent ("The concentration of the additive during the standard assay was as follows: 10 mM inhibitor, 10% detergent, and 50% (v/v) or 90% (v/v) organic solvent.")

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Ethyl Acetate

134

145

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Ethyl Acetate

129

176

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Ethyl Acetate

142

183

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Ethyl Acetate

130

151

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

1,4-Dioxane

134

161

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

1,4-Dioxane

129

178

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

1,4-Dioxane

142

180

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

1,4-Dioxane

130

152

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Acetonitrile

134

178

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Acetonitrile

129

167

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Acetonitrile

142

158

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Methanol

130

138

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

10%

134

174

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

10%

129

211

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

10%

142

180

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

10%

130

174

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

134

168

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

129

193

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

142

151

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Isopropanol

130

165

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Methanol

134

186

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Methanol

129

186

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 60°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

Activity + Stability - Incubation

Activity after incubation (5 hours at 60°C), measured by absorbance spectrophotometry (change in NADH absorbance measurement, 340 nm) in the presence of organic solvent

Methanol

142

137

Incubation: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl, Assay: 100 mM MES buffer, 5mM 2-mercaptoethanol, 100 mM KCl

Incubation: 6, Assay: 6

Incubation: 50°C, Assay: 65°C

20 mM Ethyl benzoylformate

Ethyl (R)-mandelate (95% ee)

0.3 mM NADH

—

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Visualization : Activity + Stability — Incubation

One bar per measurement. Colour = solvent, shade = solvent volume.

Structure

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Short-chain alcohol dehydrogenase from Thermus thermophilus HB27

Enzyme Description

Sequence

Experimental Data (24 measurements)

Visualization : Activity + Stability — Incubation

Structure

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