Inorganic pyrophosphatase from Haloferax volcanii

Enzyme Description

Species

Haloferax volcanii ↗

Extremophile

Yes "halophilic" organism

EC Number

3.6.1.1 ↗

Sequence

Length: 177 amino acids

MVNLWEDMETGPNAPDEIYAVVECLKGERNKYEYDKDIPGVVLDRVLHSNVHYPSDYGFIPQTYYDDEDPFDVLVLVEDQTFPGCVIEARPVALMKMDDDGEQDDKVIAVPVEDPRYDHIEDLDDIPQQTLDEIDEFFATYKNLEAGKEVETLGWEDKQAAKDAIEHAMDLYEENFA

Lana J. McMillan et al. (2016) — Archaeal Inorganic Pyrophosphatase Displays Robust Activity under High-Salt Conditions and in Organic Solvents

Applied and Environmental Microbiology · doi:10.1128/AEM.03055-15 ↗ · Activity - Classical Stability - Incubation

16 measurements ▼

Database ID

UniProt: D4GT97 ↗

Sequence Annotation

Explicit - Provided

Protein Source

Recombinant, host archaeon Haloferax volcanii H26-pJAM2920

Experimental Data (16 measurements)

16 measurements

Property	Assay	Solvent	Solvent Volume	Aqueous Reference	Measured Value	Units	Solution	pH	Temperature	Substrate(s)	Product(s)	Cofactor(s)	Shaking	Comments
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	25%	100	63	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Dimethylformamide (DMF)	25%	100	94	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Ethanol	25%	100	110	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Methanol	25%	100	150	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	30%	100	91	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	30%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	30%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	30%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	40%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	40%	100	94	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	40%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	40%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	50%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	50%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	50%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase

Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Dimethyl Sulfoxide (DMSO)	25%	100	63	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Dimethylformamide (DMF)	25%	100	94	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Ethanol	25%	100	110	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Activity - Classical	Activity measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in the presence of organic solvent	Methanol	25%	100	150	%	20 mM Tris-HCl, 1.5 M NaCl, 10 mM MgCl₂	8	Room temperature	1.5 mM Pyrophosphate	inorganic orthophosphate	—	—	Classical aqueous control (in %)
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	30%	100	91	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	30%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	30%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	30%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	40%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	40%	100	94	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	40%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	40%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Ethanol	50%	100	93	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Methanol	50%	100	96	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethylformamide (DMF)	50%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase
Stability - Incubation	Activity after incubation (2 hours on ice), measured by absorbance spectrophotometry (colorimetric assay, orthophosphate and ammonium molybdate reaction product absorbance measurement, 630 nm) in aqueous phase	Dimethyl Sulfoxide (DMSO)	50%	100	99	%	Incubation: 20 mM Tris-HCl, 2M NaCl, Assay: 20 mM Tris-HCl, 2M NaCl, 2.5 mM MgCl₂	Incubation: 8, Assay: 8	Incubation: On ice, Assay: Room temperature	50 µM Pyrophosphate	inorganic orthophosphate	—	—	Water-incubated control (in %), assay measured in aqueous phase \| Clear about assay in aqueous phase

Visualization : Activity — Classical

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

Visualization : Stability — Incubation