Sialylation of human IgG-Fc carbohydrate by transfected rat alpha 2,6-sialyltransferase

A recombinant IgG3 antibody with Phe-243 replaced by Ala (FA243) was expres sed in a CHO-K1 parental cell line. The resulting IgG-Fc-linked carbohydrat e was significantly alpha2,3-sialylated (53% of glycans), as indicated by n ormal- and reverse-phase HPLC analyses. Following transfection of a rat alp ha2,6-sialyltransferase gene into this parental cell line, IgG-Fc-linked gl ycans were sialylated (60% of glycans) such that the ratio of alpha2,6- to alpha2,3-linked sialic acid was 0.9:1.0. By comparison, the wild-type IgG3 (F243) is minimally sialylated (2-3% alpha2,3-linked), thus suggesting that sialylation is controlled primarily by the protein structure local to the carbohydrate and that the two sialyltransferases compete to sialylate the n ascent oligosaccharide. The additional alpha -2,6-sialylation affected the function of the recombinant antibody. FA243 IgG3 having both alpha2,6 and a lpha2,3-sialylation restored recognition to wild-type IgG3 levels for human Fc gamma RI, Fc gamma RII, and target cell lysis by complement. We discuss how sialylation linkage could modulate IgG function. (C) 2001 Academic Pre ss.