IDENTIFICATION AND CHARACTERIZATION OF A 230-KDA GOLGI-ASSOCIATED PROTEIN RECOGNIZED BY AUTOANTIBODIES FROM A PATIENT WITH HBV HEPATITIS

A serum from a patient with HBV hepatitis was found to contain autoant ibodies reacting with various mammalian cells, Immunofluorescence stai ning of cultured cells with the autoantibodies revealed that the antig en was localized at perinuclear regions, where the Golgi markers alpha -mannosidase II and beta-COP were colocalized, The autoantigen disappe ared from the perinuclear regions upon incubation with the fungal meta bolite brefeldin A, and the immunostainable structures were fragmented into vesicles by treatment with nocodazole. These results strongly in dicate that the antigen is localized at the Golgi complex. Immunoblots of cell lysates showed that the autoantibodies recognized a single pr otein with a molecular mass of 230 kDa in a variety of cell lines, ind icating that the 230-kDa antigen is a conserved protein among mammalia n species. We designated this protein GCP230 (Golgi complex-associated protein with a molecular mass of 230 kDa). When a post-nuclear fracti on was prepared and centrifuged, GCP230 was recovered in both cytosol and membrane fractions. Peripheral interaction of GCP230 with membrane s was confirmed by phase separation in Triton X-114 solution and by ex traction with sodium carbonate. Taken together, these results indicate that GCP230 is a peripheral membrane protein of the Golgi derived fro m the cytosol, although its function is not known at present.