Synthesis of pyridoxamine 5′-phosphate using an MBA:pyruvate transaminase as biocatalyst

Transaminases (TAs) have useful applications as biocatalysts because of their capability of introducing amino groups into ketones and keto acids with high enantioselectivity, regioselectivity and broad substrate specificity. In this study we have shown that purified His-tagged omega-TA CV2025 from Chromobacterium violaceum is capable of complete conversion of pyridoxal 5′-phosphate (PLP) to pyridoxamine 5′-phosphate (PMP) in the presence of (S)-α-methylbenzylamine (MBA) as the amine donor. Conversions of 5 mM PLP with at least 0.8 mg/ml CV2025 TA (5.8 U/ml) were complete within 24 h. The fastest completion was achieved with an enzyme concentration of 3 mg/ml (22 U/ml): Within 4 h 5 mM PLP/MBA were converted to 100% and 10 mM PLP/MBA to 70%. PLP amination was only partially inhibited in the presence of 0.5 mM gabaculine, whereas the MBA:pyruvate transamination was shown to be inhibited completely. PMP formation of comparable efficiency could not be achieved with equivalent units of porcine α-TA. This represents the first example of a PLP-converting TA with an attributed gene and the first demonstration of quantitative biocatalytic PMP synthesis.