Solubilization and properties of galactosyltransferase and sulfotransferase activities of Golgi membranes in Triton X-100.

UDP-galactose-A-acetylglucosamine galactosyltransferase of rat liver Golgi apparatus is membrane-bound, that is, it is not released by treatment with salt or disruption of the Golgi saccules using N, decompression in a Parr bomb. The enzyme can be solubilized to greater than 90% by treatment of rat liver Golgi membranes with 2 mg of Triton X-lOO/ml. We have determined the sedimentation constant (3.2 s) and the partial specific volume (0* = 0.82) of the enzyme.Triton X-100 complex by sedimentation of the solubilized Golgi membrane proteins in sucrose gradients made in water or D,O containing Triton X-100. The amount of Triton X-100 present in the complex can be calculated to be 0.52 g/g of complex if we assume that the partial specific volume of the protein portion of the enzyme.detergent complex is 0.735 and that of the detergent portion is 0.908. The Stokes’ radius of the complex was determined by chromatography of the solubilized Golgi membrane proteins on Sepharose 6B in the presence of 2 mg of Triton X-1001ml. The Stokes’ radius of the complex is 48 A and the frictional ratio (f/f,) of the complex is 1.4. The molecular weight of the complex is 97,200, of which 52% is due to bound Triton X-100. The molecular weight of the protein is therefore estimated to be 46,500. This is remarkably similar to values of 43,000 to 54,000 determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate for the three soluble galactosyltransferase isoenzymes purified from bovine milk by Barker et al. ((1972) J. Biol. Chem. 247, 7135). In contrast to the milk enzyme, however, the liver Golgi enzyme is highly lipophilic and appears to be an intrinsic membrane protein. Galactosyltransferase and 3-phosphoadenosine-5’-phosphosulfate: cerebroside sulfotransferase activities of rat kidney Golgi apparatus also are membrane-bound enzymes which can be solubilized by treatment with Triton X-100. The sedimentation constants (Q,, W) and the partial specific volumes (ti*) of the enzyme.Triton X-100 complexes have been determined. For rat kidney galactosyltransferase, s*~,~ = 4.0 and i(* = 0.85, while for the sulfotransferase, sBO,rc. = 3.6 and d* = 0.84. From these data, the weight fractions of Triton X-