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Literature summary for 2.2.1.6 extracted from

  • Steinmetz, A.; Vyazmensky, M.; Meyer, D.; Barak, Z.E.; Golbik, R.; Chipman, D.M.; Tittmann, K.
    Valine 375 and phenylalanine 109 confer affinity and specificity for pyruvate as donor substrate in acetohydroxy acid synthase isozyme II from Escherichia coli (2010), Biochemistry, 49, 5188-5199.
    View publication on PubMed

Protein Variants

Protein Variants Comment Organism
F109M both substrate affinity and kcat are significantly compromised. The specificity for 2-ketobutyrate as acceptor is not altered Escherichia coli
V375A slightly reduced kcat value with a moderate increase of the apparent KM of pyruvate. The specificity for 2-ketobutyrate as acceptor is not altered Escherichia coli
V375I slightly reduced kcat value with a moderate increase of the apparent KM of pyruvate. The specificity for 2-ketobutyrate as acceptor is not altered Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
6.6
-
pyruvate wild-type, pH 7.6, 37°C Escherichia coli
7.3
-
pyruvate mutant V375I, pH 7.6, 37°C Escherichia coli
9.1
-
2-oxobutanoate mutant V375A, pH 7.6, 37°C Escherichia coli
13.8
-
pyruvate mutant V375A, pH 7.6, 37°C Escherichia coli
17.3
-
pyruvate mutant F109M, pH 7.6, 37°C Escherichia coli
300
-
2-oxobutanoate wild-type, pH 7.6, 37°C Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
acetohydroxy acid synthase isozyme II
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.4
-
mutant F109M, pH 7.6, 37°C Escherichia coli
8.2
-
mutant V375A, pH 7.6, 37°C Escherichia coli
22.7
-
mutant V375I, pH 7.6, 37°C Escherichia coli
34.3
-
wild-type, pH 7.6, 37°C Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2 pyruvate 60fold higher specificity for 2-ketobutyrate over pyruvate as acceptor Escherichia coli 2-acetolactate + CO2
-
?
2-oxobutanoate + pyruvate 60fold higher specificity for 2-ketobutyrate over pyruvate as acceptor Escherichia coli 2-hydroxy-2-methyl-3-oxopentanoate + CO2
-
?
additional information a valine and a phenylalanine residue hydrophobically interact with the methyl substituent of pyruvate. A mutation of either Val375 or Phe109 is detrimental for unimolecular catalytic steps in which tetrahedral intermediates are involved, such as substrate addition to the cofactor and product liberation. Val375 and Phe109 to not only conjointly mediate substrate binding and specificity but moreover to ensure a proper orientation of the donor substrate and intermediates for correct orbital alignment in multiple transition states Escherichia coli ?
-
?

Synonyms

Synonyms Comment Organism
AHAS
-
Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.3
-
2-oxobutanoate wild-type, pH 7.6, 37°C Escherichia coli
3.9
-
pyruvate mutant F109M, pH 7.6, 37°C Escherichia coli
9.6
-
pyruvate mutant V375A, pH 7.6, 37°C Escherichia coli
11.9
-
2-oxobutanoate mutant V375A, pH 7.6, 37°C Escherichia coli
26.6
-
pyruvate mutant V375I, pH 7.6, 37°C Escherichia coli
40.3
-
pyruvate wild-type, pH 7.6, 37°C Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.011
-
2-oxobutanoate wild-type, pH 7.6, 37°C Escherichia coli
0.23
-
pyruvate mutant F109M, pH 7.6, 37°C Escherichia coli
0.7
-
pyruvate mutant V375A, pH 7.6, 37°C Escherichia coli
1.3
-
2-oxobutanoate mutant V375A, pH 7.6, 37°C Escherichia coli
3.6
-
pyruvate mutant V375I, pH 7.6, 37°C Escherichia coli
6.1
-
pyruvate wild-type, pH 7.6, 37°C Escherichia coli