XYLOGLUCAN GLUCOSYLTRANSFERASE IN GOLGI MEMBRANES FROM PISUM-SATIVUM (PEA)

Cell membranes from etiolated Pisum sativum (pea) tissues were separat ed by ultracentrifugation on linear sucrose density gradients and assa yed for membrane marker and glycosyltransferase activity. Membrane fra ctions were shown to incorporate glucose from UDP-D-[C-14]glucose into polysaccharides with glycosyl linkages consistent with synthesis of x yloglucan. A combined assay using g.c., radiogas proportional counting and m.s. was employed to determine the identities of C-14-labelled gl ycosyl residues and the glycosyl linkages between them. In glucan synt hase I assays, membrane fractions enriched for Golgi membranes showed C-14 incorporation into 4- and 4,6-glucose residues, with minor incorp oration into 3-glucose residues. In glucan synthase II assays, all C-1 4 incorporation was into 3- and 3,4-glucose. There was a shift in glyc osyl linkage of C-14 incorporation from predominantly 4-glucose at low UDP-glucose concentration to predominantly 3- and 3,4-glucose at high UDP-glucose concentrations. Mn2+ stimulated incorporation of radioact ivity into 4,6-glucose residues characteristic of xyloglucan polysacch arides. Addition of exogenous UDP-xylose to assay mixtures stimulated incorporation into 4,6-glucose, with a maximum at 15 muM UDP-xylose.