BIOSYNTHESIS OF HEPARIN .25. SUBSTRATE SPECIFICITIES OF GLYCOSYLTRANSFERASES INVOLVED IN FORMATION OF HEPARIN PRECURSOR AND ESCHERICHIA-COLI K5 CAPSULAR POLYSACCHARIDES

The E. coli K5 capsular polysaccharide is composed of 4)GlcpA(beta 1-4 )GlcpNAc(alpha 1-disaccharide units. A partially N-deacetylated/N-sulf ated heptasaccharide, derived from this polymer and having a nonreduci ng terminal GlcNAc unit, was used as acceptor for a mastocytoma micros omal GlcA-transferase involved in heparin biosynthesis. An octasacchar ide with nonreducing-terminal GlcA similarly served as acceptor for th e microsomal GlcNAc-transferase. Analysis of the labeled octa- and non a-saccharides formed by transfer of monosaccharide units from UDP-[C-1 4]GlcA and UDP-[H-3]GlcNAc, respectively, showed that both glycosyltra nsferases could utilize partially N-sulfated accepters. The GlcA-trans ferase showed a marked preference for a terminal GlcNAc-GlcA-GlcNSO(3) -sequence, particularly when this sequence was followed by an addition al N-sulfated disaccharide unit. Enzymes catalyzing the same GlcA and GlcNAc transfer reactions were solubilized from E. coli K5 membranes. The K5 capsular polysaccharide, like the heparin/heparan sulfate precu rsor polysaccharide, thus probably grows by stepwise, alternating addi tion of the two constituent monosaccharide units, from the correspondi ng UDP-sugars, to the nonreducing ends of the chains. Moreover, the ba cterial glycosyltransferases utilized the same partially N-sulfated ol igosaccharide substrates as the mammalian enzymes, and with similar pr eference for N-sulfate groups in certain positions.