SBV333-ATA FROM STREPTOMYCES SP. BV333: FROM DISCOVERY TO ENGINEERING

Aminotransferases (ATAs) are pyridoxal-5'-phosphate (PLP)-dependent enzymes capable of catalyzing the synthesis of chiral amines, making them highly valuable for various industrial applications. Through a combination of genome mining and functional screening, a putative transaminase gene from Streptomyces sp. BV333 was identified, showing sequence similarity to known thermostable ATAs.[1] The corresponding enzyme, Sbv333-ATA, was successfully expressed in E. coli and exhibited remarkable thermostability (Tm = 85 °C) along with broad substrate specificity toward various amino acceptors.[2] To further investigate its substrate scope, a panel of amine donors was also tested. Notably, the enzyme displayed good activity toward several amine donors, including (S)-methylbenzylamine and 2-phenylethylamine, as well as aliphatic mono- and diamines such as propylamine and cadaverine, in addition to selected amino acids. However, more sterically hindered aromatic amines were not accepted as substrates by this biocatalyst. The enzyme was successfully crystallized, and high-resolution structures were obtained for both its native form and its complex with the inhibitor gabaculine. Based on its three-dimensional structure, a rational mutagenesis approach was employed to expand the substrate scope of Sbv333-ATA. Several variants were successfully overexpressed in E. coli and among these, the W89A variant showed the highest activity toward bulky amine substrates, such as the diaromatic compound 1,2-diphenylethylamine. High-resolution structures of the most promising Sbv333-ATA variants were also determined.[3]