Topology of the Maize Mixed Linkage (1(right arrow)3),(1(right arrow)4)-(beta)-D-Glucan Synthase at the Golgi Membrane

Mixed-linkage (1(right arrow)3),(1(right arrow)4)-(beta)-D-glucan is a plant cell wall polysaccharide composed of cellotriosyl and cellotetraosyl units, with decreasingly smaller amounts of cellopentosyl, cellohexosyl, and higher cellodextrin units, each connected by single (1(right arrow)3)-(beta)-linkages. (1(right arrow)3),(1(right arrow)4)-(beta)-Glucan is synthesized in vitro with isolated maize (Zea mays) Golgi membranes and UDP-[14C]D-glucose. The (1(right arrow)3),(1(right arrow)4)-(beta)-glucan synthase is sensitive to proteinase K digestion, indicating that part of the catalytic domain is exposed to the cytoplasmic face of the Golgi membrane. The detergent {3-[(3cholamidopropyl)dimethylammonio]-1-propanesulfonic acid} (CHAPS) also lowers (1(right arrow)3),(1(right arrow)4)-(beta)glucan synthase activity. In each instance, the treatments selectively inhibit formation of the cellotriosyl units, whereas synthesis of the cellotetraosyl units is essentially unaffected. Synthesis of the cellotriosyl units is recovered when a CHAPS-soluble factor is permitted to associate with Golgi membranes at synthesis-enhancing CHAPS concentrations but lost if the CHAPS-soluble fraction is replaced by fresh CHAPS buffer. In contrast to other known Golgiassociated synthases, (1(right arrow)3),(1(right arrow)4)-(beta)-glucan synthase behaves as a topologic equivalent of cellulose synthase, where the substrate UDP-glucose is consumed at the cytosolic side of the Golgi membrane, and the glucan product is extruded through the membrane into the lumen. We propose that a cellulose synthase-like core catalytic domain of the (1(right arrow)3),(1(right arrow)4)-(beta)-glucan synthase synthesizes cellotetraosyl units and higher even-numbered oligomeric units and that a separate glycosyl transferase, sensitive to proteinase digestion and detergent extraction, associates with it to add the glucosyl residues that complete the cellotriosyl and higher oddnumbered units, and this association is necessary to drive polymer elongation.