Sallie O. Hoch

Antigenic domains on the U1 small nuclear ribonucleoprotein-associated 70K polypeptide: A comparison of regions selectively recognized by human and mouse autoantibodies and by monoclonal antibodies

Antigenic regions on the U1 small nuclear ribonucleoprotein (snRNP)-associated 70K polypeptide recognized by human and mouse autoantibodies or by monoclonal antibodies were identified and compared. Using a set of 70K fusion proteins as antigen in enzyme-linked immunosorbent assay and immunoblotting revealed that serum autoantibodies of human and of MRL/Mp mouse origin recognized a common region of the 70K polypeptide. Monoclonal anti-70K antibodies derived from a patient with mixed connective tissue disease, from an autoimmune MRL/Mp mouse, and from a BALB/c mouse immunized with purified U1 snRNP were all shown to bind to a part of the 70K polypeptide rich in charged residues and different from the region recognized by most human and MRL/Mp mouse serum autoantibodies.

Mapping of the immunoreactive domains of a small nuclear ribonucleoprotein-associated Sm-D autoantigen

The Sm-D(D1) small nuclear ribonucleoprotein (snRNP) polypeptide is a major target of autoantibodies diagnostic for systemic lupus erythematosus. The cDNA encoding the protein from Raji cells was expressed in Escherichia coli as a fusion protein with anthranilate synthase (TrpE-Sm-D). When tested by protein blot, the recombinant polypeptide was strongly immunoreactive under defined blotting conditions, which appear to facilitate the refolding of the polypeptide into a native conformation. Multiple translational fusions between the trpE gene and fragments encompassing the length of the Sm-D coding sequence were constructed for epitope mapping. The results describe two general patterns of anti-Sm reactivity: (i) antibodies that recognize only the full-length antigen and are presumably directed against discontinuous epitopes, and (ii) antibodies that recognize the carboxy terminus of the antigen which embodies an extended/charged structure.