2026-09-22T12:30:08Zhttps://repository.ajou.ac.kr/oai/request

oai:repository.ajou.ac.kr:201003/43582024-10-11T02:17:39Zcom_201003_14372com_201003_14368col_201003_14373

C-terminus of Hepatitis B virus RNase H Domain is Important for HBV replication B형 간염 바이러스 DNA중합효소 RNase H domain 의 C-말단이 바이러스 증식에 미치는 영향 김, 태영 김, 경민 대학원 의생명과학과 200924464 김, 태영 B형 간염 바이러스 DNA 중합효소 RNase H Domain C-말단 C-Terminus Hepatitis B Virus HBV replication Hepatitis B Virus (HBV) DNA polymerase (P) protein consisting of terminal protein (TP), spacer, reverse transcriptase (RT), and RNase H, plays critical roles in viral assembly and replication. RNase H domain is required for HBV DNA replication, however critical motif or amino acid residues in the RNase H domain for the HBV replication has not been extrensively demonstrated yet. In the present study, several chimeras of P protein by substituting Duck hepatitis B virus (DHBV) sequences were constructed. Accordingly, we tested a series of P protein chimeras in which several substitution mutants were disigned to contain various amino acids of DHBV P protein. It is found that amino acid residues from 800 to 826 (800SRPLLRLPFQPTTGRTSLYAVSPSVPS826) in C -terminus of the RNase H domain are required to complete HBV replication. HBV P protein mutants in which single amino acid residue was substituted were examined for the rescue of HBV replication. Among these mutants tested, L806T mutant P protein have a defect in pgRNA encapsidation and viral DNA synthesis, demonstrating that leucine at position 806 is critical for HBV replication. Ⅰ. INTRODUCTION 1 Ⅱ. MATERIALS AND METHODS 5 A. HBV plasmid DNA construction 5 B. Cell culture and transfection 8 C. Isolation of core particles 9 D. RNase protection assay (RPA) 9 E. Core particle Western blotting 10 F. Southern blotting 11 G. SDS-PAGE and Western blotting 11 Ⅲ. RESULT 12 A. HBV P constructs containing DHBV P residues in RNase H domain 12 B. Amino acid residues from 800 to 826 in C-terminus of the RNase H are critical for pgRNA encapsidaion and DNA synthesis. 14 C. The small motif substituted HBV RNase H domain mutants at C-terminus have ability to support HBV DNA synthesis. 21 D. The small motif substituted HBV RNase H domain mutants at C-terminus have ability to support HBV pgRNA encapsidation. 23 E. A leucine residue at position 806 in HBV P protein is important for viral genome replication. 26 F. A leucine residue at position 806 in HBV P protein is important for pgRNA encapsidation. 28 Ⅳ. DISCUSSION 29 Ⅴ. CONCLUSION 31 REFERENCES 32 국문요약 38 Master 2011-11-07T02:17:00Z 2011-11-07T02:17:00Z 2011 2011 Thesis Theses http://repository.ajou.ac.kr/handle/201003/4358 http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000011459 000000011459 en