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Detailed Structure–Function Correlations of Bacillus subtilis Acetolactate Synthase
Author(s) -
Sommer Bettina,
von Moeller Holger,
Haack Martina,
Qoura Farah,
Langner Clemens,
Bourenkov Gleb,
Garbe Daniel,
Loll Bernhard,
Brück Thomas
Publication year - 2015
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201402541
Subject(s) - isobutanol , bacillus subtilis , acetolactate synthase , isobutyraldehyde , thiamine pyrophosphate , cofactor , stereochemistry , chemistry , active site , enzyme , biochemistry , biology , alcohol , genetics , catalysis , bacteria
Isobutanol is deemed to be a next‐generation biofuel and a renewable platform chemical.1 Non‐natural biosynthetic pathways for isobutanol production have been implemented in cell‐based and in vitro systems with Bacillus subtilis acetolactate synthase (AlsS) as key biocatalyst.2–6 AlsS catalyzes the condensation of two pyruvate molecules to acetolactate with thiamine diphosphate and Mg 2+ as cofactors. AlsS also catalyzes the conversion of 2‐ketoisovalerate into isobutyraldehyde, the immediate precursor of isobutanol. Our phylogenetic analysis suggests that the ALS enzyme family forms a distinct subgroup of ThDP‐dependent enzymes. To unravel catalytically relevant structure‐function relationships, we solved the AlsS crystal structure at 2.3 Å in the presence of ThDP, Mg 2+ and in a transition state with a 2‐lactyl moiety bound to ThDP. We supplemented our structural data by point mutations in the active site to identify catalytically important residues.