The 1.2-kb DNA fragment of the Rickettsia conorii outer membrane protein B gene (OmpB 451-846 ) was subcloned using site-specific PCR primers and expressed as six smaller fragments: OmpB 458-652 , OmpB 595-744 , OmpB 595-654 , OmpB 645-692 , OmpB 689-744 , and OmpB 739-848 . NCTC cells transfected with a mammalian expression vector expressing the fragments OmpB 689-744 and OmpB 739-848 stimulated immune anti- R . conorii CD8 T lymphocytes, suggesting the presence of CD8 T-lymphocyte-stimulating epitopes on these fragments. In order to further characterize the CD8 T-lymphocyte-stimulatory elements, CD8 T-lymphocyte epitopes on OmpB 689-744 and OmpB 739-848 were mapped by overlapping synthetic peptides. The ability of these synthetic peptides to stimulate immune CD8 T lymphocytes was determined by gamma interferon (IFN-γ) production and cell proliferation after incubation with simian virus 40-transformed murine vascular endothelial cells in the presence of a 20 μM solution of each synthetic peptide. Five synthetic peptides, SKGVNVDTV (OmpB 708-716 ), ANVGSFVFN (OmpB 735-743 ), IVSGTVGGQ (OmpB 749-757 ), ANSTLQIGG (OmpB 789-797 ), and IVEFVNTGP (OmpB 812-820 ), induced secretion of IFN-γ at significantly higher levels than the controls. Three of these five peptides, SKGVNVDTV (OmpB 708-716 ), ANSTLQIGG (OmpB 789-797 ), and IVEFVNTGP (OmpB 812-820 ), also stimulated the proliferation of immune CD8 T lymphocytes. Significantly higher levels of specific cytotoxic T-lymphocyte killing were observed with the same three synthetic peptides, SKGVNVDTV (OmpB 708-716 ), ANSTLQIGG (OmpB 789-797 ), and IVEFVNTGP (OmpB 812-820 ).
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