MAMMALIAN GLYCOSYLTRANSFERASES PREFER GLYCOSYL PHOSPHORYL DOLICHOLS RATHER THAN GLYCOSYL PHOSPHORYL POLYPRENOLS AS SUBSTRATES FOR OLIGOSACCHARYL SYNTHESIS

We have studied the effectiveness of polyprenyl-P-mannose and polypren ol-P-glucose as donor substrates for the dolichyl-P-mannose:Man(5)(Glc NAc)(2)-PP-dolichol mannosyltransferase and the dolichyl-P-glucose: Ma n(9)(GlcNAc)(2)-PP-dolichol glucosyltransferase, respectively. The pol yprenol moiety differs from dolichol only in the unsaturation of the t erminal isoprene unit of the molecule. Based on the kinetics of the re actions, we have found that both glycosyltransferases have higher appa rent K(m)s and lower apparent V(max)s using polyprenyl-P-monosaccharid es as substrates rather than the dolichyl-P-monosaccharides The produc ts formed with the polyprenyl-P-sugars were the same as those formed b y the dolichol-linked sugars, indicating that the polyprenol substrate s could be utilized by the glycosyltransferases in vitro. The results also indicate that the dolichyl-P-sugars and the polyprenyl-P-sugars c ompete for the same binding site on the enzyme. These findings are sig nificant in terms of understanding the glycosylation phenotypes of Chi nese hamster ovary cell mutants of the Lec9 complementation group, whi ch lack the ability to convert polyprenol into dolichol. (C) 1994 Acad emic Press, Inc.