Purification and Characterization of Oligomeric Species of an Aromatic Amino Acid Aminotransferase From Lactococcus Lactis Subsp. Lactis S3

Two oligomeric species of an aromatic amino acid aminotransferase (Ar-AT) were purified and characterized from Lactococcus lactis subsp. lactis S3. The more abundant species, Ar-AT1, was purified over 2,200-fold with a 3% recovery. The molecular masses of Ar-AT1 were determined to be 42 kDa and 84 kDa by SDS-PAGE and gel filtration, respectively. The molecular masses of the less abundant species, Ar-AT2, were 42 kDa and 170 kDa, respectively. Both Ar-AT1 and Ar-AT2 have identical N-terminal sequences which indicates they are homodimeric and homotetrameric forms of the same enzyme. The Ar-ATs catalyze pyridoxal-5′-phosphate dependent transamination of Phe, Tyr, Trp, Leu and Met utilizing α-ketoglutarate as the amino acceptor. Km values of the two Ar-ATs for Trp, Tyr and Phe ranged from 0.65 to 2.43 mM. However, differences were observed in the pI and specific activities between the two Ar-ATs. The pI values of Ar-AT1 and Ar-AT2 were 4.63 and 3.93, respectively. The Vmax/Km ratios of Ar-AT2 for Trp, Tyr and Phe were three-fold greater than that of Ar-AT1, indicating that Ar-AT2 is more catalytically efficient on these amino acids than Ar-AT1.