J Biol Chem, Vol. 274, Issue 26, 18206-18212, June 25, 1999

On the Biosynthesis of Alternating -2,9/-2,8 Heteropolymer of Sialic Acid Catalyzed by the Sialyltransferase of Escherichia coli Bos-12

From the Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan, Republic of China

Escherichia coli Bos-12 synthesizes a heteropolymer of sialic acids with alternating -2,9/-2,8 glycosidic linkages (1). In this study, we have shown that the polysialyltransferase of the E. coli Bos-12 recognizes an -2,8 glycosidic linkage of sialic acid at the nonreducing end of an exogenous acceptor of either the -2,8 homopolymer of sialic acid or the alternating -2,9/-2,8 heteropolymer of sialic acid and catalyzes the transfer of Neu5Ac from CMP-Neu5Ac to this residue. When the exogenous acceptor is an -2,8-linked oligomer of sialic acid, the main product synthesized is derived from the addition of a single residue of [¹⁴C]Neu5Ac to form either an -2,8 glycosidic linkage or an -2,9 glycosidic linkage at the nonreducing end, at an -2,8/-2,9 ratio of approximately 2:1. When the acceptor is the alternating -2,9/-2,8 heteropolymer of sialic acid, chain elongation takes place four to five times more efficiently than the -2,8-linked homopolymer of sialic acid as an acceptor. It was found that the -2,9-linked homopolymer of sialic acid and the -2,8/-2,9-linked hetero-oligomer of sialic acid with -2,9 at the nonreducing end not only failed to serve as an acceptor for the E. coli Bos-12 polysialyltransferase for the transfer of [¹⁴C]Neu5Ac, but they inhibited the de novo synthesis of polysialic acid catalyzed by this enzyme. The results obtained in this study favor the proposal that the biosynthesis of the -2,9/-2,8 heteropolymer of sialic acid catalyzed by the E. coli Bos-12 polysialyltransferase involves a successive transfer of a preformed -2,8-linked dimer of sialic acid at the nonreducing terminus of the acceptor to form an -2,9 glycosidic linkage between the incoming dimer and the acceptor. The glycosidic linkage at the nonreducing end of the alternating -2,9/-2,8 heteropolymer of sialic acid produced by E. coli Bos-12 should be an -2,8 glycosidic bond and not an -2,9 glycosidic linkage.