Glycobiology, published online 9 January 2013; doi:10.1093/glycob/cwt003

Capsular polysaccharides—sugar chains that appear on the surface of many bacteria—vary in both the carbohydrates used and the linkages between these residues. Escherichia coli K1 features a polysaccharide chain composed of α2,8-linked sialic acids, whereas that of the related E. coli K92 consists of sialic acids with alternating α2,8 and α2,9 linkages. However, each strain uses only a single enzyme, a polysialyltransferase, to construct the chains. To understand how the polysialyltransferase from K92 creates alternating linkages, Keys et al. first created a mutant library varying the six residues in the K92 transferase that are different from the K1 sequence within an N-terminal region previously identified as relevant for enzyme specificity. The mutants were transformed into an engineered E. coli BL21 strain, in which the capsule biosynthesis genes are dormant, and the resultant polysaccharides were evaluated using linkage-specific antibodies and chemical characterization. The authors observed the formation of α2,8-linked structures but not α2,9-linked sequences, with each successful functional switch linked to a H52N mutation. Introduction of the inverse N52H mutation into the K1 sequence similarly switched its specificity to that of K92's enzyme. The authors speculate that the residue controls the positioning of the polysaccharide acceptor, placing either the C8 or C9 hydroxyl group of the terminal sugar closer to the incoming donor sugar and thus determining the reaction outcome.