One-pot enzymatic synthesis of UDP-D-glucose from UMP and glucose-1-phosphate using an ATP regeneration system

Abstract

Glucose-1-phosphate uridylyltransferase from E. coli K12 was used to convert uridine-5′-triphosphate and glucose-1-phosphate to UDP-D-glucose. The conversion was efficient and completed within 5 minutes under the employed conditions. In addition, thymidine-5′-monophosphate kinase and acetate kinase were proven to be non-specific, converting udridine-5′- monophosphate to uridine-5′-triphosphate with 55% conversion after 6h, which was much slower than the production of TTP under the same conditions (complete conversion within one hour). Since these two reactions could proceed under the same conditions, a one-pot synthesis of UDP-D-glucose with ATP regeneration was designed from easily available starting materials, and conversion up to 40% by HPLC peak integration was achieved given a reaction time of 4 h.