Strand transfer is enhanced by mismatched nucleotides at the 3' primer terminus: a possible link between HIV reverse transcriptase fidelity and recombination.

摘要

Strand transfer catalyzed by HIV reverse transcriptase (RT) was examined. The system consisted of a 142 nt RNA (donor) to which a 50 nt DNA primer was hybridized. The primer bound such that its 3' terminal nucleotide hybridized to the 12th nt from the 5' end of the donor. The 3' terminal nucleotide on the primer was either a G, A or T residue. Since the corresponding nucleotide of the donor was a C, the G formed a matched terminus and the A or T a mismatched terminus. The efficiency with which DNA bound to the donor transferred to a second RNA, termed acceptor, was monitored. The acceptor was homologous to the donor for all but the last 9 nt at the 5' end of the donor. Therefore, homologous strand transfer could occur at any point prior to the DNA being extended into the nonhomologous region on the donor. Strand transfer occurred approximately twice as efficiently with the mismatched versus matched substrates. The mismatched nucleotide was fixed into transfer products indicating that excision of the mismatch was not required for RT extension or transfer. Results suggest that base misincorporations by RT may promote recombination by enhancing strand transfer.