Purification and Characterization of α-Acetolactate Decarboxylase from Brevibacterium acetylicum

Microorganisms capable of producing high amounts of α-acetolactate decarboxylase (ALDC; EC 4.1.1.5) were screened for with stock type cultures. Brevibacterium acetylicum had the most potent enzyme activity among the strains tested. The productivity of ALDC by B. acetylicum was elevated by adding Zn2+ to the medium. ALDC was purified from the cell-free extract of B. acetylicum by a procedure involving ammonium sulfate fractionation, Sephadex G-100 gel filtration, and DEAE-cellulose and FPLC-MonoQ column chromatographies. The purified enzyme was homogeneous by polyacrylamide gel electrophoresis. The molecular weight of the native enzyme was 62,000 by TSK-gel filtration and the subunit molecular weight was 31,000 by SDS polyacrylamide gel electrophoresis. The enzyme activity was inhibited by metal chelators such as diethyldithiocarbamate, 8-oxyquinoline, and o-phenanthroline. Analysis by atomic absorption spectrophotometry showed that zinc atoms were involved in the purified enzyme preparation.