Roles of the Human Immunodeficiency Virus Type 1 Nucleocapsid Protein in Annealing and Initiation versus Elongation in Reverse Transcription of Viral Negative-Strand Strong-Stop DNA
AUTOR(ES)
Rong, Liwei
FONTE
American Society for Microbiology
RESUMO
To study the initiation of human immunodeficiency virus type 1 reverse transcription, we have used the viral nucleocapsid protein (NC7) to anneal tRNA3Lys primer onto viral genomic RNA and have then eliminated NC7 from this primer-template complex by digestion with proteinase K and phenol-chloroform extraction of residual protein. Our data show that saturating concentrations of NC7 resulted in the formation of an active tRNA-template complex that yielded enhanced production of full-length negative-strand strong-stop DNA [(−)ssDNA] and that this complex remained active even after the elimination of NC7. While both of the two Zn finger motifs found within NC7 were essential for efficient elongation, NC protein that contained a point mutation in the first Zn finger or that was devoid of both Zn fingers yielded primer-template complexes that could still be initiated in 1-base-extension assays. In contrast, the use of heat annealing to produce primer-template complexes resulted in proportions of full-length (−)ssDNA lower than those seen with NC protein, and the addition of NC protein to such preformed primer-template complexes was able to reverse this defect only to a marginal extent.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=110360Documentos Relacionados
- Human immunodeficiency virus type 1 nucleocapsid protein promotes efficient strand transfer and specific viral DNA synthesis by inhibiting TAR-dependent self-priming from minus-strand strong-stop DNA.
- In Vitro Analysis of Human Immunodeficiency Virus Type 1 Minus-Strand Strong-Stop DNA Synthesis and Genomic RNA Processing
- Plus-strand strong-stop DNA synthesis in retrotransposon Ty1.
- Effects of 3'-deoxynucleoside 5'-triphosphate concentrations on chain termination by nucleoside analogs during human immunodeficiency virus type 1 reverse transcription of minus-strand strong-stop DNA.
- Premature strand transfer by the HIV-1 reverse transcriptase during strong-stop DNA synthesis.