| Summary: | 碩士 === 國立中興大學 === 生物化學研究所 === 97 === The influenza A virus non-structural protein 1 (NS1) is a multifunctional protein and a virulence factor. It is considered to antagonize host innate immune responses. However, there is no clear correlation between the function of NS1 and its virulence. NS1 structure can be divided into two distinct functional domains. An N-terminal double-stranded RNA binding domain (RBD), which binds double-strand RNA and inhibit the activity of RNase L system and a C-terminal effector domain. NS1 has been shown to associate with several cellular proteins via the effector domain, including CPSF 30, a cellular factor required for 3´ end processing of cellular pre-mRNA, thereby inhibiting the production of all cellular mRNAs. As NS1 can dimerize and is coupled with dsRNA binding activity, we want to understand the correlation between its dimeric structure and functions by constructing several NS1 mutants disrupting the NS1 dimeric interface. We then examine the dimerization activity of NS1 mutants via GST pulldown assay and bacteria two hybrid assay, we use EMSA technique to analyze the dsRNA binding activity. Unexpectedly, we found the S103F/I106M/R35A/R46A mutant represses interferon-beta expression stronger than the S103F/I106M mutant, although the dimerization and dsRNA binding activity of S103F/I106M/R35A/R46A are impaired severely. Therefore, it should be cautious in the attempt of disrupting NS1 dimeric interface to decrease its virulence toward host cell.
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