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Journal of Virology, January 2004, p. 1012-1019, Vol. 78, No. 2
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.2.1012-1019.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Increased G->A Transition Frequencies Displayed by Primer Grip Mutants of Human Immunodeficiency Virus Type 1 Reverse Transcriptase

Clara E. Cases-González and Luis Menéndez-Arias*

Centro de Biología Molecular "Severo Ochoa," Consejo Superior de Investigaciones Científicas—Universidad Autónoma de Madrid, Cantoblanco, Madrid, Spain

Received 25 July 2003/ Accepted 26 September 2003

A genetic screen based on the blue-white ß-galactosidase complementation assay designed to detect G->A mutations arising during RNA-dependent DNA synthesis was used to compare the fidelity of mutant human immunodeficiency virus type 1 reverse transcriptases (RTs) with the mutations M230L and M230I with the wild-type enzyme, in the presence of biased deoxynucleoside triphosphate (dNTP) pools. The mutant RTs with the M230L and M230I changes were found to be 20 to 70 times less faithful than the wild-type RT in the presence of low [dCTP]/[dTTP] ratios but showed similar fidelity in assays carried out with equimolar concentrations of each nucleotide. Biased dNTP pools led to short tandem repeat deletions in the target sequence, which were also detectable with the assay. However, deletion frequencies were similar for all of the RTs tested. The reported data suggest that RT pausing due to the low dNTP levels available in the RT reaction mixture facilitates strand transfer, in a process that is not necessarily mediated by nucleotide misinsertion.

* Corresponding author. Mailing address: Centro de Biología Molecular "Severo Ochoa," CSIC-Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain. Phone: 34-914978477. Fax: 34-914974799. E-mail: lmenendez{at}cbm.uam.es.

Journal of Virology, January 2004, p. 1012-1019, Vol. 78, No. 2
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.2.1012-1019.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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