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Literature summary extracted from

  • Eram, M.; Sarafuddin, B.; Gong, F.; Ma, K.
    Characterization of acetohydroxyacid synthase from the hyperthermophilic bacterium (2015), Biochem. Biophys. Rep., 4, 89-97.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.2.1.6 expressed in Escherichia coli BL21 (DE3) Rosetta 2 cells Thermotoga maritima
2.2.1.6 expression in Escherichia coli Thermotoga maritima
2.2.1.6 recombinant expression of the catalytic subunit in Escherichia coli strain BL21 (DE3) Rosetta 2, subcloning in Escherichia coli strain DH5alpha, reconstitution of the catalytic and regulatory subunits leads to increased AHAS activity Thermotoga maritima
2.2.1.6 recombinant expression of the regulatory subunit in Escherichia coli strain BL21 (DE3) Rosetta 2, subcloning in Escherichia coli strain DH5alpha, reconstitution of the catalytic and regulatory subunits leads to increased AHAS activity Thermotoga maritima

General Stability

EC Number General Stability Organism
2.2.1.6 stirring has detrimental effects on activity. When stirred, both aerobic and anaerobic aliquots lose major portions of their activity at almost the same rate resulting in a time required for the loss of its 50% activity of about 18 h Thermotoga maritima

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.2.1.6 additional information
-
 additional information Michaelis-Menten steady-state kinetic analysis, overview Thermotoga maritima
2.2.1.6 additional information
-
 additional information activity is dependent on the ionic strength of the buffer and diminishes considerably (approximately 80%) when assayed in buffers with less than 100 mM concentrations. At concentrations higher than 100 mM the activity levels are quite similar (tested up to 500 mM) Thermotoga maritima
2.2.1.6 16.4
-
 pyruvate pH 7.0, 80°C Thermotoga maritima
2.2.1.6 16.4
-
 pyruvate pH 7.0, 85°C, recombinant catalytic subunit Thermotoga maritima
2.2.1.6 16.4
-
 pyruvate at pH 7.0 and 80°C Thermotoga maritima

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.2.1.6 Mg2+ required Thermotoga maritima
2.2.1.6 additional information the AHAS activity is dependent on the ionic strength of the buffer and diminishes considerably (approximately 80%) when assayed in buffers with less than 100 mM concentrations. At concentrations higher than 100 mM the activity levels are quite similar (up to 500 mM is tested), suggesting it might be needed to maintain the functional oligomeric state of the enzyme Thermotoga maritima

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
2.2.1.6 20508
-
 2 * 20508, regulatory subunit, sequence calculation, 2 * 22210, regulatory subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 20508
-
 2 * 65497 + 2 * 20508, calculated from amino acid sequence Thermotoga maritima
2.2.1.6 22200
-
 1 * 65497, calculated, 1 * 66400, SDS-PAGE, plus 1 * 2058, calculated, 1 * 22200, SDS-PAGE, catalytic and regulatory subunit, respectively Thermotoga maritima
2.2.1.6 22200
-
 2 * 66400 + 2 * 22200, SDS-PAGE Thermotoga maritima
2.2.1.6 22210
-
 2 * 20508, regulatory subunit, sequence calculation, 2 * 22210, regulatory subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 37700
-
 recombinant catalytic subunit, gel filtration Thermotoga maritima
2.2.1.6 65497
-
 2 * 65497, catalytic subunit, sequence calculation, 2 * 66398, catalytic subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 65497
-
 1 * 65497, calculated, 1 * 66400, SDS-PAGE, plus 1 * 2058, calculated, 1 * 22200, SDS-PAGE, catalytic and regulatory subunit, respectively Thermotoga maritima
2.2.1.6 65497
-
 2 * 65497 + 2 * 20508, calculated from amino acid sequence Thermotoga maritima
2.2.1.6 66398
-
 2 * 65497, catalytic subunit, sequence calculation, 2 * 66398, catalytic subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 66400
-
 1 * 65497, calculated, 1 * 66400, SDS-PAGE, plus 1 * 2058, calculated, 1 * 22200, SDS-PAGE, catalytic and regulatory subunit, respectively Thermotoga maritima
2.2.1.6 66400
-
 2 * 66400 + 2 * 22200, SDS-PAGE Thermotoga maritima
2.2.1.6 156000
-
 recombinant catalytic subunit, gel filtration Thermotoga maritima
2.2.1.6 156400
-
 gel filtration Thermotoga maritima
2.2.1.6 156800
-
 gel filtration Thermotoga maritima

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.2.1.6 2 pyruvate Thermotoga maritima
-
 2-acetolactate + CO2
-
 ?
2.2.1.6 pyruvate + 2-oxobutyrate Thermotoga maritima
-
 2-aceto-2-hydroxybutyrate + CO2
-
 ?
2.2.1.6 pyruvate + pyruvate Thermotoga maritima
-
 2-acetolactate + CO2
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.2.1.6 Thermotoga maritima
-
-
-
2.2.1.6 Thermotoga maritima Q9WZ19 small subunit
-
2.2.1.6 Thermotoga maritima Q9WZ19 and Q9WZ18 Q9WZ19 i.e. regulatory subunit, Q9WZ18 i.e. catalytic subunit
-
2.2.1.6 Thermotoga maritima DSM 3109 Q9WZ19 and Q9WZ18 Q9WZ19 i.e. regulatory subunit, Q9WZ18 i.e. catalytic subunit
-

Oxidation Stability

EC Number Oxidation Stability Organism
2.2.1.6 enzyme is not oxygen sensitive Thermotoga maritima

Purification (Commentary)

EC Number Purification (Comment) Organism
2.2.1.6
-
 Thermotoga maritima
2.2.1.6 DEAE-Sepharose column chromatography and hydroxyapatite column chromatography Thermotoga maritima
2.2.1.6 recombinant catalytic and regulatory subunits from Escherichia coli strain BL21(DE3) Rosetta 2, the catalytic subunit is purified 30.2fold by heat treatement, and anion exchange and hydroxyapatite chromatography Thermotoga maritima
2.2.1.6 recombinant regulatory subunit from Escherichia coli strain BL21(DE3) Rosetta 2, the regulatory subunit is purified to homogeneity by heat treatment at 80°C for 1h and nickel affinity chromatography Thermotoga maritima

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.2.1.6 5.4
-
 crude extract, at pH 7.0 and 80°C Thermotoga maritima
2.2.1.6 163
-
 pH 7.0, 85°C, purified recombinant catalytic subunit Thermotoga maritima
2.2.1.6 163
-
 after 30.2fold purification, at pH 7.0 and 80°C Thermotoga maritima

Storage Stability

EC Number Storage Stability Organism
2.2.1.6 0°C, purified recombinant enzyme, 5 days, 35-40% loss of activity Thermotoga maritima

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.2.1.6 2 pyruvate
-
 Thermotoga maritima 2-acetolactate + CO2
-
 ?
2.2.1.6 2 pyruvate
-
 Thermotoga maritima DSM 3109 2-acetolactate + CO2
-
 ?
2.2.1.6 pyruvate + 2-oxobutyrate
-
 Thermotoga maritima 2-aceto-2-hydroxybutyrate + CO2
-
 ?
2.2.1.6 pyruvate + pyruvate
-
 Thermotoga maritima 2-acetolactate + CO2
-
 ?

Subunits

EC Number Subunits Comment Organism
2.2.1.6 dimer 2 * 20508, regulatory subunit, sequence calculation, 2 * 22210, regulatory subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 dimer 2 * 65497, catalytic subunit, sequence calculation, 2 * 66398, catalytic subunit, SDS-PAGE Thermotoga maritima
2.2.1.6 dimer 1 * 65497, calculated, 1 * 66400, SDS-PAGE, plus 1 * 2058, calculated, 1 * 22200, SDS-PAGE, catalytic and regulatory subunit, respectively Thermotoga maritima
2.2.1.6 heterotetramer 2 * 65497 + 2 * 20508, calculated from amino acid sequence Thermotoga maritima
2.2.1.6 heterotetramer 2 * 66400 + 2 * 22200, SDS-PAGE Thermotoga maritima
2.2.1.6 More the enzyme consists of a catalytic and a regulatory subunit Thermotoga maritima

Synonyms

EC Number Synonyms Comment Organism
2.2.1.6 acetohydroxy acid synthase
-
 Thermotoga maritima
2.2.1.6 acetohydroxyacid synthase
-
 Thermotoga maritima
2.2.1.6 AHAS
-
 Thermotoga maritima
2.2.1.6 AHASL
-
 Thermotoga maritima
2.2.1.6 AHASS
-
 Thermotoga maritima
2.2.1.6 TM_0548
-
 Thermotoga maritima
2.2.1.6 TM_0549
-
 Thermotoga maritima

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.2.1.6 85
-
-
 Thermotoga maritima

Temperature Range [°C]

EC Number Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
2.2.1.6 additional information
-
 incubation at either 70 or 80°C results in enhanced activity with the highest activity achieved after 1 h of incubation, longer treatments result in reduced activity Thermotoga maritima
2.2.1.6 50 100 activity range, overview Thermotoga maritima

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.2.1.6 98
-
 pyruvate pH 7.0, 80°C Thermotoga maritima
2.2.1.6 98
-
 pyruvate pH 7.0, 85°C, recombinant catalytic subunit Thermotoga maritima
2.2.1.6 98
-
 pyruvate at pH 7.0 and 80°C Thermotoga maritima

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.2.1.6 7
-
-
 Thermotoga maritima

pH Range

EC Number pH Minimum pH Maximum Comment Organism
2.2.1.6 4.5 10.5 activity range, overview Thermotoga maritima
2.2.1.6 5.5 8 more than 50% between pH 5.5 and 8.0 Thermotoga maritima

Cofactor

EC Number Cofactor Comment Organism Structure
2.2.1.6 FAD
-
 Thermotoga maritima
2.2.1.6 FAD required Thermotoga maritima
2.2.1.6 FAD apparent Km value 0.00015 mM Thermotoga maritima
2.2.1.6 thiamine diphosphate
-
 Thermotoga maritima
2.2.1.6 thiamine diphosphate dependent on Thermotoga maritima
2.2.1.6 thiamine diphosphate apparent Km value 0.057 mM Thermotoga maritima

General Information

EC Number General Information Comment Organism
2.2.1.6 metabolism acetohydroxyacid synthase is the key enzyme in branched chain amino acid biosynthesis pathway, overview Thermotoga maritima