Abstract
The topological disposition of a form of UDP-glucuronyltransferase (called GT-1) in rat liver microsomes was examined. Concanavalin A-Sepharose failed to bind microsomal vesicles even though GT-1 has sugar chains of “high mannose” type, indicating that mannose-containing sugar chains of microsomal glycoproteins including GT-1 are not exposed to the outer surface of microsomal vesicles. Polyclonal antibodies raised against purified GT-1 could bind to microsomal vesicles, indicating that at least part of the GT-1 polypeptide chain is extruded to the outside of the microsomal membrane. Intact microsomal vesicles were digested with carboxypeptidase Y and then subjected to immunoblot analysis using the anti-GT-1 antibodies. It was thus found that the digestion resulted in cleavage of a C-terminal, 2-kDa fragment, leaving a 52-kDa fragment of GT-1 still tightly bound to the membrane. From these results, it is concluded that GT-1 is a transmembrane protein, which extrudes its C-terminal end (at least 2 kDa) to the outside of the membrane, whereas most of its polypeptide chain together with the sugar chains are located on the luminal side of the membrane.
