title: Molecular cloning and expression of Rickettsia tsutsugamushi genes for two major protein antigens in Escherichia coli.
abstract: Several polypeptide antigens of Rickettsia tsutsugamushi are recognized by human or primate convalescent sera and may be important protective immunogens. Molecular cloning and expression of the genes encoding the 110K (110 kilodalton) and 56K polypeptide antigens of R. tsutsugamushi Karp were accomplished in the lambda gt11 expression vector system. Southern blot analysis with the cloned fragments for the 56K polypeptide antigen (0.7 kilobases) and the 110K polypeptide antigen (5.4 kilobases) confirmed that the insert DNA was rickettsial and not host cell in origin. Expression of a complete 110K polypeptide was shown to be independent of isopropyl-beta-D-thiogalactopyranoside induction, suggesting that an intact rickettsial promoter was operational. Epitopes of the 56K polypeptide were expressed as lac promoter-dependent beta-galactosidase fusion proteins. Polyclonal antibody, affinity purified against the recombinant 110K and 56K polypeptides, reacted with polypeptides of similar size in the Kato and Gilliam strains of R. tsutsugamushi. Group-reactive, but not strain-specific, monoclonal antibodies against the 56K polypeptide reacted with the cloned portion of the 56K polypeptide. Western blot analysis demonstrated that the cloned 56K Karp antigen gene product is recognized by human convalescent serum.
mesh_terms: D000907:Antibodies, Bacterial / Q000276:immunology; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000235:genetics* / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D006801:Humans; D008970:Molecular Weight; D012285:Orientia tsutsugamushi / Q000235:genetics* / Q000276:immunology; D012612:Scrub Typhus / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D001426:Bacterial Proteins