Substitution of proline 306 in the reverse transcriptase domain of hepatitis B virus regulates replication.
The proline residue at position 306 in hepatitis B virus (HBV) reverse transcriptase (rtP306) has been suggested to constrain the conformation of the alpha-helices in the thumb subdomain that interacts with the viral DNA template-primer. To study the impact of residue rt306 in HBV replication further, 11 site-directed mutants were constructed that substituted rtP306 with different amino acids. The replicative competencies of these mutants were assayed by HepG2 cell transfection and real-time PCR. When rtP306 was substituted with glycine or threonine, the replication competency of these mutants was drastically reduced to 1.96 and 4.51% of the wild-type HBV level, respectively. When rtP306 was substituted with glutamic acid, the replicative competency of the mutant increased up to 9.4-fold compared with wild-type virus. The results also showed that changes in the replicative competency of these constructed mutants were not associated with functional changes of HBV enhancer I. These results indicate the importance of amino acid(s) at the interface between the thumb and palm subdomains in modulating the replicative competency of HBV isolates. This regulatory residue(s) could serve as a new target for the development of anti-HBV drugs.
Lin
Xu
X
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
Ma
Zhang-Mei
ZM
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
Yao
Xin
X
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
He
Li-Fang
LF
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
Yuan
Zheng-Hong
ZH
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
Ding
Jian-Ping
JP
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, PR China.
Wen
Yu-Mei
YM
Key Laboratory of Medical Molecular Virology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.
D019943:Amino Acid Substitution; D002460:Cell Line; D006515:Hepatitis B virus / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D011392:Proline / Q000737:chemistry / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014779:Virus Replication
D011392:Proline; D012194:RNA-Directed DNA Polymerase