Novel Approach to the Biosynthesis of a Potentially Therapeutic Immunomodulatory Bacterial Polysaccharide

Abstract

Capsular Polysaccharide A (CPSA) is found on the surface of Bacteroides fragilis, a symbiotic gut microbe. CPSA is made up of repeats of a tetrasaccharide subunit. Our laboratory is applying an in vitro method to enzymatically synthesize CPSA, which is assembled on a bactoprenyl diphosphate scaffold one sugar at a time. There is evidence suggesting that the first sugar incorporated into this scaffold is acetamido‐4‐amino‐6‐deoxy‐glactopryanose (AADGal). I isolated the presumed dehydratase, UngD2, aminotransferase, WcfR, and the initiating glycosyltransferase, WcfS, proposed to be responsible for bactoprenyl‐dipohsophatelinked AADGal biosynthesis. Utilizing capillary electrophoresis, ion exchange high performance liquid chromatography (HPLC), reverse phase HPLC, and ultraviolet light active bactoprenyl monophosphate analogues we were able to characterize the function of WcfR and WcfS. The role of UngD2, however, appeared to be ambiguous in the conditions tested thus far.