GM3 trisaccharide biosynthesis and process optimization using engineered E. coli lysate and whole-cell catalysis

An engineered E. coli strain BL21 (DE3, SiaB, PmST3, △lacZ, △nanZ) that simultaneously encoded CMP-Neu5Ac synthase (SiaB) and α-2-3-sialyltransferase (PmST3) was constructed to synthesize GM3 trisaccharide. The crude E. coli lysate could be readily used for biosynthesis of GM3 trisaccharide with a nearly quantitative yield under optimal conditions. In addition, this engineered E. coli strain as whole-cell catalyst could generate approximately 1 g of GM3 trisaccharide in 40 mL of medium under optimal conditions. The whole-cell catalyst could be recycled and reused for at least 3 rounds without obvious loss of enzymatic activity.