Halohydrin dehalogenase (gene HHDH) from Agrobacterium radiobacter AD1

Enzyme Description

Species

Extremophile

EC Number

4.5.1.1 ↗

Sequence

Length: 254 amino acids

MSTAIVTNVKHFGGMGSALRLSEAGHTVACHDESFKQKDELEAFAETYPQLKPMSEQEPAELIEAVTSAYGQVDVLVSNDIFAPEFQPIDKYAVEDYRGAVEALQIRPFALVNAVASQMKKRKSGHIIFITSATPFGPWKELSTYTSARAGACTLANALSKELGEYNIPVFAIGPNYLHSEDSPYFYPTEPWKTNPEHVAHVKKVTALQRLGTQKELGELVAFLASGSCDYLTGQVFWLAGGFPMIERWPGMPE

Nevena Milčić et al. (2022) — Inhibitory Effect of DMSO on Halohydrin Dehalogenase: Experimental and Computational Insights into the Influence of an Organic Co-solvent on the Structural and Catalytic Properties of a Biocatalyst

Chemistry–A European Journal · doi:10.1002/chem.202201923 ↗ · Activity - Classical Stability - deltaHTm Stability - Inactivation Rate Constant Stability - Incubation Stability - Tm Stability - Half-life

73 measurements ▼

Database ID

UniProt: Q93D82 ↗

Sequence Annotation

Explicit - Provided PDB Accession Number

Protein Source

Recombinant, host bacterium Escherichia coli MC1061

Experimental Data (73 measurements)

73 measurements — page 4 of 4

Property	Assay	Solvent	Solvent Volume	Aqueous Reference	Measured Value	Units	Solution	pH	Temperature	Substrate(s)	Product(s)	Cofactor(s)	Shaking	Comments
Stability - Incubation	Activity after incubation (76 hours at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	40%	100.0	24.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (0.1 hour at 50°C),measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	65.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (1 hour at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	29.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (2.5 hours at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	18.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (4 hours at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	12.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (29 hours at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	2.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (72 hours at 50°C), measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100.0	1.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Incubation	Activity after incubation (4 hours at 50°C),measured by HPLC in aqueous phase	Dimethyl Sulfoxide (DMSO)	30%	100.0	94.0	%	Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer	Incubation: 7.5, Assay: 7.5	Incubation: 25°C, Assay: 25°C	5 mM p-Nitro-2-bromo-1-phenylethanol	para-nitro styrene oxide	—	Incubation: 1000 rpm	Non-incubated control (in %), assay in aqueous phase \| Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements
Stability - Tm	Tm measured by DSC in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	50%	60.0	37.7	°C	100 mM Tris-SO4 buffer	7.5	—	—	—	—	—	Classical aqueous control (in °C) \| Enzyme cell-free extracts used for measurements
Stability - Tm	Tm measured by DSC in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	40%	60.0	43.2	°C	100 mM Tris-SO4 buffer	7.5	—	—	—	—	—	Classical aqueous control (in °C) \| Enzyme cell-free extracts used for measurements
Stability - Tm	Tm measured by DSC in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	30%	60.0	46.9	°C	100 mM Tris-SO4 buffer	7.5	—	—	—	—	—	Classical aqueous control (in °C) \| Enzyme cell-free extracts used for measurements
Stability - Tm	Tm measured by DSC in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	20%	60.0	54.1	°C	100 mM Tris-SO4 buffer	7.5	—	—	—	—	—	Classical aqueous control (in °C) \| Enzyme cell-free extracts used for measurements
Stability - Tm	Tm measured by DSC in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	5%	60.0	58.6	°C	100 mM Tris-SO4 buffer	7.5	—	—	—	—	—	Classical aqueous control (in °C) \| Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

35%

4.7

0.5

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

2 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

5.2

2.9

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) with 1% organic solvent | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

5.2

1.1

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) with 1% organic solvent | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

35%

5.2

0.7

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) with 1% organic solvent | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

5.2

0.4

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) with 1% organic solvent | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethylformamide (DMF)

20%

100.0

73.0

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in %) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethylformamide (DMF)

35%

100.0

22.0

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in %) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethylformamide (DMF)

50%

100.0

4.0

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in %) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

4.7

0.3

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

2 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

10%

5.2

1.9

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) with 1% organic solvent | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

4.7

0.8

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

2 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

10%

4.7

1.4

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

2 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

4.7

2.5

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

2 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

4.2

0.3

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

1 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

35%

4.2

0.4

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

1 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

4.2

0.5

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

1 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

10%

4.2

1.0

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

1 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

4.2

1.9

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

1 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

5.2

0.4

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

4 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

35%

5.2

0.7

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

4 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

5.2

1.1

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

4 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

10%

5.2

1.9

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

4 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

5.2

3.0

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

4 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

2.8

1.6

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

3 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

10%

2.8

0.9

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

3 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

2.8

0.6

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

3 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

2.8

0.3

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

3 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Activity - Classical

Activity measured by HPLC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

35%

2.8

0.3

U/mg

100 mM Tris-SO4 buffer

7.5

25°C

3 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in U/mg) | Enzyme cell-free extracts used for measurements

Stability - ΔH_Tm

δ HTM measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

40%

509.0

358.0

kJ/mol

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in kJ/mol) | Enzyme cell-free extracts used for measurements

Stability - ΔH_Tm

δ HTM measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

509.0

517.0

kJ/mol

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in kJ/mol) | Enzyme cell-free extracts used for measurements

Stability - ΔH_Tm

δ HTM measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

509.0

502.0

kJ/mol

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in kJ/mol) | Enzyme cell-free extracts used for measurements

Stability - ΔH_Tm

δ HTM measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

30%

509.0

513.0

kJ/mol

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in kJ/mol) | Enzyme cell-free extracts used for measurements

Stability - ΔH_Tm

δ HTM measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

509.0

175.0

kJ/mol

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in kJ/mol) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethylformamide (DMF)

50%

520.0

0.51

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethylformamide (DMF)

35%

520.0

0.57

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethylformamide (DMF)

20%

520.0

1.0

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

50%

520.0

0.7

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

40%

520.0

41.0

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Half-life

Half-life (in hours), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

30%

520.0

484.0

hours

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in hours) | Enzyme cell-free extracts used for measurements

Stability - Inactivation Rate Constant

Deactivation constant (in 1/min), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

40%

2.22*10^-5

2.81*10^-4

1/min

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in 1/min) | Enzyme cell-free extracts used for measurements

Stability - Inactivation Rate Constant

Deactivation constant (in 1/min), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

30%

2.22*10^-5

2.39*10^-5

1/min

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in 1/min) | Enzyme cell-free extracts used for measurements

Stability - Inactivation Rate Constant

Deactivation constant (in 1/min), determined by residual activity measurements (HPLC) after incubation at 25°C

Dimethyl Sulfoxide (DMSO)

50%

2.22*10^-5

1.67*10^-2

1/min

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Classical aqueous control (in 1/min) | Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (52 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

20%

100.0

90.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 25°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

30%

97.0

96.5

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Water-incubated control (in %) | Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 25°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

97.0

67.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Water-incubated control (in %) | Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 25°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

97.0

0.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Water-incubated control (in %) | Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (4 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

100.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

100.0

95.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (48 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

100.0

95.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (72 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

100.0

89.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (2 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

20%

100.0

97.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (29 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

20%

100.0

98.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (48 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

30%

100.0

91.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (72 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

20%

100.0

93.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

30%

100.0

92.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (76 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

30%

100.0

89.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (2 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

100.0

93.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (5 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

100.0

90.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (24 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

100.0

66.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (48 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

100.0

46.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (76 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

40%

100.0

24.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (0.1 hour at 50°C),measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

65.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (1 hour at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

29.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (2.5 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

18.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (4 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

12.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (29 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

2.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (72 hours at 50°C), measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

50%

100.0

1.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Incubation

Activity after incubation (4 hours at 50°C),measured by HPLC in aqueous phase

Dimethyl Sulfoxide (DMSO)

30%

100.0

94.0

Incubation: 100 mM Tris-SO4 buffer, Assay: 100 mM Tris-SO4 buffer

Incubation: 7.5, Assay: 7.5

Incubation: 25°C, Assay: 25°C

5 mM p-Nitro-2-bromo-1-phenylethanol

para-nitro styrene oxide

—

Incubation: 1000 rpm

Non-incubated control (in %), assay in aqueous phase | Uncertainty about whether the assay really was in aqueous phase/Enzyme cell-free extracts used for measurements

Stability - Tm

Tm measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

50%

60.0

37.7

°C

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in °C) | Enzyme cell-free extracts used for measurements

Stability - Tm

Tm measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

40%

60.0

43.2

°C

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in °C) | Enzyme cell-free extracts used for measurements

Stability - Tm

Tm measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

30%

60.0

46.9

°C

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in °C) | Enzyme cell-free extracts used for measurements

Stability - Tm

Tm measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

20%

60.0

54.1

°C

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in °C) | Enzyme cell-free extracts used for measurements

Stability - Tm

Tm measured by DSC in the presence of organic solvent

Dimethyl Sulfoxide (DMSO)

60.0

58.6

°C

100 mM Tris-SO4 buffer

7.5

—

Classical aqueous control (in °C) | Enzyme cell-free extracts used for measurements

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Visualization : Activity — Classical

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

Visualization : Stability — T_m

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

Visualization : Stability — ΔH_Tm

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

Visualization : Stability — Incubation

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

Visualization : Stability — Half-life

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

Visualization : Stability — Inactivation Rate Constant

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

Structure

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Halohydrin dehalogenase (gene HHDH) from Agrobacterium radiobacter AD1

Enzyme Description

Sequence

Experimental Data (73 measurements)

Visualization : Activity — Classical

Visualization : Stability — Tm

Visualization : Stability — ΔHTm

Visualization : Stability — Incubation

Visualization : Stability — Half-life

Visualization : Stability — Inactivation Rate Constant

Structure

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Visualization : Stability — T_m

Visualization : Stability — ΔH_Tm