The linear mapping for B cell epitope on HCMV pp65 antigen by Systemic Lupus Erythematosus patients' sera

• Main Authors: 許佑民許佑民許佑民許佑民許佑民, 許佑民
• Other Authors: Chang Mingi
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/40933005226323771875

碩士 === 慈濟大學 === 分子生物及細胞生物研究所 === 94 === Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease. It causes multiple organ damage and presents a variety of clinical and laboratory phenomena, particularly inflammation. Clinically, SLE is often accompanied by different autoantibodies, such as anti-double strand DNA Ab (anti-dsDNA Ab), anti-Smith Ab (anti-Sm Ab), anti-snRNP Ab, anti-Ro/La Ab (anti-SSa/SSb Ab) and ect. Current investigations agreed that the mechanisms involved in promoting SLE including genetics (such as human leukocyte antigen (HLA) types)、complement deficiency、MHC、sex hormone、environmental factor (such as drug)、virus infection、ultraviolet light and stress. Viruses are considered possible aetiologic agents of autoimmune disease. There have been reported that human cytomegalovirus (HCMV) may be a pathogenetic factor in systemic lupus erythematosus (SLE). We have reported pp65 antigen, which is a 65 kDa phosphoprotein encoded by HCMV, was recognized by SLE patients’ sera. Following immunization in female NZB/W F1 mice, it induced early onset of autoimmunity. Those results indicated that SLE patients have strong B cell activity against the HCMV pp65 antigen. Partial mapping the associated B cell epitope in pp65 antigen found that only the pp65-3, that cover one third of the entire pp65 on C terminal, was recognized by SLE patients’sera. For the subsequent fine mapping of B cell epitope within pp65-3, the pp65-3 fragment was divided into pp65-336-379, pp65-378-455 and pp65-454-561 fragments respectively. Following expressing and purifications, fragments were detected by SLE patients’sera via immunoblotting and ELISA assay. The results shown that SLE patients’sera recognize pp65-378-455 at a higher frequency than pp65-336-379 and pp65-454-561. Subsequently, to pin point the reactive epitope, we constructed pp65-336-448, pp65-336-439 and pp65-336-422 fragments. Immunoblotting and ELISA results revealed decreased positive frequencies by SLE patients’sera between pp65-336-439 and pp65-336-422. According to these results, studies suggested that one possible B cell epitope may locate in pp65-422-423 region that could play a role in SLE development.