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Functional Role of Hepatitis B Virus Core Protein in Viral Replication
- Alternative Title
- 정재성
- Author(s)
- Jung, Jaesung
- Alternative Author(s)
- 정재성
- Advisor
- 김경민
- Department
- 일반대학원 의생명과학과
- Publisher
- The Graduate School, Ajou University
- Publication Year
- 2013-02
- Language
- eng
- Keyword
- Hepatits B Virus
- Alternative Abstract
- -ABSTRACT- Functional Role of Hepatitis B Virus Core Protein in Viral Replication PART I Arginine-Rich 167RRRSQSPRR175 Domain in C-Terminus of Core is Critical for HBV Replication To investigate the contributions of carboxyl-terminal nucleic acid binding domain of HBV core protein for hepatitis B virus (HBV) replication, chimeric HBV core proteins were generated by substituting varying lengths of the carboxyl-terminus of duck hepatitis B virus (DHBV) core protein for the corresponding regions of HBV core protein. All chimeric core proteins formed core particles. A chimeric core protein with 221–262 amino acids of DHBV core protein, in place of 146–185 amino acids of the HBV core protein, supported HBV pregenomic RNA (pgRNA) encapsidation and DNA synthesis: 40% amino acid sequence identity or 45% homology in the nucleic-acid binding domain of HBV core protein was sufficient for pgRNA encapsidation and DNA synthesis, although we predominantly detected spliced DNA. A chimeric core protein with 221–241 and 251–262 amino acids of DHBV core, in place of HBV C 146–166 and 176–185 amino acids, respectively, could rescue full-length DNA synthesis. However, a reciprocal core chimera with 242–250 of DHBV core (242RAGSPLPRS250) introduced in place of 167–175 of HBV core (167RRRSQSPRR175) significantly decreased pgRNA encapsidation and DNA synthesis, and full-length DNA was not detected, demonstrating that the arginine-rich 167RRRSQSPRR175 domain may be critical for efficient viral replication. Five amino acids differing between viral species (underlined above) were tested for replication rescue; R169 and R175 were found to be important. PART II Phosphorylation of the C-Terminal Domain of HBV Core Protein Modulates Genome Replication Phosphorylation of hepatitis B virus (HBV) core protein at Ser157, Ser164, and Ser172 residues by host serine/arginine protein-specific kinases (SRPK) or protein kinase C (PKC) has been demonstrated to modulate HBV replication. Also, three additional amino acid residues, Thr162, Ser170, and Ser178, of HBV core protein have been suggested as the putative protein kinase A (PKA) phosphorylation sites with the conserved RRXS/T motif. The in vivo phosphorylaiton assay reveals that Thr 162, Ser170, or Ser178 can be phosphorylated. In order to elucidate importance of these residues for HBV replication, each was mutated to Ala to mimic nonphosphorylated Ser or to Glu to mimic phosphorylated Ser. Thr 162 to Ala (T162A) mutation decreased replicative intermediate DNA significantly. To further investigate the importance of Thr 162 in conjunction with Ser170Ala and/or Ser178Ala mutations, more core protein mutants were constructed. In the presence of T162A mutation, the HBV DNA synthesis was decreased more dramatically, indicating that Thr 162 residue may be important for HBV DNA synthesis. Taken together, our results indicate that the putative PKA phosphorylation sites, Thr 162, Ser170, or Ser178, is phosphorylated and can modulate DNA replication possibly through phosphorylation and dephosphorylation.
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- Appears in Collections:
- Graduate School of Ajou University > Department of Biomedical Sciences > 3. Theses(Master)
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