Incorporation of deoxyribonucleotides and ribonucleotides by a dNTP-binding cleft mutated reverse transcriptase in hepatitis B virus core particles.
Our recent observation that hepatitis B virus (HBV) DNA polymerase (P) might initiate minus-strand DNA synthesis without primer [Kim et al., (2004) Virology 322, 22-30], raised a possibility that HBV P protein may have the potential to function as an RNA polymerase. Thus, we mutated Phe 436, a bulky amino acid with aromatic side chain, at the putative dNTP-binding cleft in reverse transcriptase (RT) domain of P protein to smaller amino acids (Gly or Val), and examined RNA polymerase activity. HBV core particles containing RT dNTP-binding cleft mutant P protein were able to incorporate (32)P-ribonucleotides, but not HBV core particles containing wild type (wt), priming-deficient mutant, or RT-deficient mutant P proteins. Since all the experiments were conducted with core particles isolated from transfected cells, our results indicate that the HBV RT mutant core particles containing RT dNTP-binding cleft mutant P protein could incorporate both deoxyribonucleotides and ribonucleotides in replicating systems.
Kim
Hee-Young
HY
Department of Microbiology, Ajou University School of Medicine, Woncheon-dong 5, Suwon 443-721, South Korea.
Kim
Hye-Young
HY
Jung
Jaesung
J
Park
Sun
S
Shin
Ho-Joon
HJ
Kim
Kyongmin
K
D045744:Cell Line, Tumor; D003854:Deoxyribonucleotides / Q000235:genetics / Q000378:metabolism*; D015684:Gene Products, pol / Q000235:genetics / Q000378:metabolism; D006515:Hepatitis B virus / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D009154:Mutation; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012265:Ribonucleotides / Q000235:genetics / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*
D003854:Deoxyribonucleotides; D015684:Gene Products, pol; C066027:P protein, Hepatitis B virus; D012265:Ribonucleotides; D012194:RNA-Directed DNA Polymerase