Characterization and role of the branched-chain aminotransferase (BcaT) isolated from Lactococcus lactis subsp cremoris NCDO 763

In Lactococcus lactis, which is widely used as a starter in the cheese indu stry, the first step of aromatic and branched-chain amino acid degradation is a transamination which is catalyzed by two major aminotransferases. We h ave previously purified and characterized biochemically and genetically the aromatic aminotransferase, AraT. In the present study, we purified and stu died the second enzyme, the branched-chain aminotransferase, BcaT. We clone d and sequenced the corresponding gene and used a mutant, along with the lu ciferase gene as the reporter, to study the role of the enzyme in amino aci d metabolism and to reveal the regulation of gene transcription. BcaT catal yzes transamination of the three branched-chain amino acids and methionine and belongs to class IV of the pyridoxal S'-phosphate-dependent aminotransf erases, In contrast to most of the previously described bacterial BcaTs, wh ich are hexameric, this enzyme is homodimeric. It is responsible for 90% of the total isoleucine and valine aminotransferase activity of the cell and for 50 and 40% of the activity towards leucine and methionine, respectively . The original role of BcaT was probably biosynthetic since expression of i ts gene was repressed by free amino acids and especially by isoleucine. How ever, in dairy strains, which are auxotrophic for branched-chain amino acid s, BcaT functions only as a catabolic enzyme that initiates the conversion of major aroma precursors. Since this enzyme is still active under cheese-r ipening conditions, it certainly plays a major role in cheese flavor develo pment.