The Role of Buffer, Pyridoxal 5’‐Phosphate and Light on the Stability of the Silicibacter Pomeroyi Transaminase

Transaminases are pyridoxal 5’‐phosphate (PLP)‐dependent enzymes that transfer amino‐functions. The transaminase from Silicibacter pomeroyi (SpATA) exhibits a broad substrate spectrum. In this work we examined the effect of different conditions (light, buffer and PLP‐concentration) on the stability of SpATA, as well as the causes for these effects. The enzyme was stored either in TRIS or CHES with 0–10 mM added PLP at 22 °C. The samples were either kept dark or they were exposed to light. The results show that invariably, all samples kept in darkness exhibited longer half‐life times than the ones exposed to light. An increase in the half‐life from 8 h to 720 h could be achieved solely by keeping the sample dark. Especially samples in CHES buffer inactivated faster in light the more PLP was present, due to the degradation of PLP. In TRIS however, an imine‐bond between TRIS and PLP protects PLP from degradation. Silicibacter Pomeroyi Transaminase: The operational stability of the Silicibacter pomeroyi transaminase was evaluated and the influence of light, buffer and cofactor‐concentrations were assessed. Under ideal conditions, which contained high concentrations of cofactor and light exclusion, a half‐life of over 700 h was achieved, which decreased to only 8 h upon exposure to light.