Improving the Activity of MTSase from Sulfolobus acidocaldarius by Error-Prone PCR

Abstract:

A mutant D-6 of MTSase from /Sulfolobus acidocaldarius ATCC 33909 was obtained through directed evolution using error-prone PCR and high-throughput screening. Gene analysis showed that there were two mutations G432D/G586D with the G586D on the loop of Aβ8 and AEα14. The enzymatic properties of wild-type MTSase and mutant D-6 was determined. The results showed that the specific activity of mutant D-6 was 1.22 times that of wild-type MTSase. The /K_m value of the mutant D-6 was 2.77 mmol/L while that of wild-type was 4.74 mmol/L. Thus, the substrate affinity of mutant increased compared to the wild-type.