B형 간염 바이러스 DNA중합효소 RNase H domain 의 C-말단이 바이러스 증식에 미치는 영향
C-terminus of Hepatitis B Virus RNase H Domain is Important for HBV replication
- 주제(키워드) HBV
- 발행기관 아주대학교
- 지도교수 김경민
- 발행년도 2011
- 학위수여년월 2011. 2
- 학위명 석사
- 학과 및 전공 일반대학원 의생명과학과
- 실제URI http://www.dcollection.net/handler/ajou/000000011459
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
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.
more목차
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
