| Summary: | 碩士 === 長庚大學 === 醫學生物技術研究所 === 97 === PB2 is a viral protein of influenza A virus. The position 627 of PB2 is considered as a determinant of host range restriction. In general, avian influenza contains glutamic acid and human influenza contains lysine at the position. Other positions on PB2 which can distinguish avian and human strains were identified by full-genome sequence analysis. We analyzed both avian and human H1N1 PB2 sequences since 1918. Position 199, 475, and 627 on PB2 are conserved evolutionally as human-like signatures in human influenza. To understand the potential roles of these 3 positions on vRNP functions, reverse genetics system based on the backbone of human influenza A/WSN/33 (H1N1) was used to address the mechanism. Mutant PB2 by single or multiple mutations on the 3 positions were generated. The double avian-like mutant, 199A+475L PB2 reduced RNP activity obviously as the single avian-like mutant, 627E PB2. Immunoprecipitation results showed that the bonding affinity of 627E PB2 to NP was weaker than that of wild type. The binding affinity of 199A+475L PB2 to NP was weaker in the presence of viral-like RNA， but it was stronger in the absence of viral-like RNA, when comparing with wild type. Accordingly, the position 199 and 475 on PB2 could be the crucial determinants to affect vRNP activity through the mechanisms different from the position 627.
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