The influence of 3TC-resistance mutations E89G and M184V in the human immunodeficiency virus reverse transcriptase on mispair extension efficiency

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Abstract

Two nucleoside analog resistance mutations in HIV-1 reverse transcriptase (RT), E89G and M184V, were previously shown to increase the dNTP insertion fidelity of HIV-1 RT. However, forward mutation assays using a lacZα reporter gene have revealed a lack of impact on the overall error rate of these variants. In an effort to investigate the basis for this discrepancy, we have examined whether the increases in misinsertion fidelity observed for E89G and M184V RTs are accompanied by an increase in mispair extension fidelity. The relative efficiencies with which the wild type, E89G, M184V and M184V/E89G HIV-1 RTs extend model template-primer duplexes containing 3'-OH terminal mismatches were measured. The calculated efficiencies of mispair extension (f(ext)) were, in general, not significantly decreased from the wild type HIV-1 RT. In fact, the efficiency of extension from one of the mispaired primer-template duplexes was significantly increased for two of the mutants tested. These results suggest that amino acid substitutions that increase the fidelity of dNTP insertion do not necessarily increase misextension fidelity, and that the decreased misextension fidelity may counterbalance the increases in misinsertion fidelity observed for E89G and M184V RTs.

Original languageEnglish (US)
Pages (from-to)4389-4394
Number of pages6
JournalNucleic Acids Research
Volume26
Issue number19
DOIs
StatePublished - Oct 1 1998

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HIV Reverse Transcriptase
Lamivudine
Mutation
Lac Operon
Amino Acid Substitution
Reporter Genes
Nucleosides
HIV-1
Human immunodeficiency virus 1 reverse transcriptase

ASJC Scopus subject areas

  • Genetics

Cite this

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title = "The influence of 3TC-resistance mutations E89G and M184V in the human immunodeficiency virus reverse transcriptase on mispair extension efficiency",
abstract = "Two nucleoside analog resistance mutations in HIV-1 reverse transcriptase (RT), E89G and M184V, were previously shown to increase the dNTP insertion fidelity of HIV-1 RT. However, forward mutation assays using a lacZα reporter gene have revealed a lack of impact on the overall error rate of these variants. In an effort to investigate the basis for this discrepancy, we have examined whether the increases in misinsertion fidelity observed for E89G and M184V RTs are accompanied by an increase in mispair extension fidelity. The relative efficiencies with which the wild type, E89G, M184V and M184V/E89G HIV-1 RTs extend model template-primer duplexes containing 3'-OH terminal mismatches were measured. The calculated efficiencies of mispair extension (f(ext)) were, in general, not significantly decreased from the wild type HIV-1 RT. In fact, the efficiency of extension from one of the mispaired primer-template duplexes was significantly increased for two of the mutants tested. These results suggest that amino acid substitutions that increase the fidelity of dNTP insertion do not necessarily increase misextension fidelity, and that the decreased misextension fidelity may counterbalance the increases in misinsertion fidelity observed for E89G and M184V RTs.",
author = "Hamburgh, {Monica E.} and Drosopoulos, {William C.} and Prasad, {Vinayaka R.}",
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T1 - The influence of 3TC-resistance mutations E89G and M184V in the human immunodeficiency virus reverse transcriptase on mispair extension efficiency

AU - Hamburgh, Monica E.

AU - Drosopoulos, William C.

AU - Prasad, Vinayaka R.

PY - 1998/10/1

Y1 - 1998/10/1

N2 - Two nucleoside analog resistance mutations in HIV-1 reverse transcriptase (RT), E89G and M184V, were previously shown to increase the dNTP insertion fidelity of HIV-1 RT. However, forward mutation assays using a lacZα reporter gene have revealed a lack of impact on the overall error rate of these variants. In an effort to investigate the basis for this discrepancy, we have examined whether the increases in misinsertion fidelity observed for E89G and M184V RTs are accompanied by an increase in mispair extension fidelity. The relative efficiencies with which the wild type, E89G, M184V and M184V/E89G HIV-1 RTs extend model template-primer duplexes containing 3'-OH terminal mismatches were measured. The calculated efficiencies of mispair extension (f(ext)) were, in general, not significantly decreased from the wild type HIV-1 RT. In fact, the efficiency of extension from one of the mispaired primer-template duplexes was significantly increased for two of the mutants tested. These results suggest that amino acid substitutions that increase the fidelity of dNTP insertion do not necessarily increase misextension fidelity, and that the decreased misextension fidelity may counterbalance the increases in misinsertion fidelity observed for E89G and M184V RTs.

AB - Two nucleoside analog resistance mutations in HIV-1 reverse transcriptase (RT), E89G and M184V, were previously shown to increase the dNTP insertion fidelity of HIV-1 RT. However, forward mutation assays using a lacZα reporter gene have revealed a lack of impact on the overall error rate of these variants. In an effort to investigate the basis for this discrepancy, we have examined whether the increases in misinsertion fidelity observed for E89G and M184V RTs are accompanied by an increase in mispair extension fidelity. The relative efficiencies with which the wild type, E89G, M184V and M184V/E89G HIV-1 RTs extend model template-primer duplexes containing 3'-OH terminal mismatches were measured. The calculated efficiencies of mispair extension (f(ext)) were, in general, not significantly decreased from the wild type HIV-1 RT. In fact, the efficiency of extension from one of the mispaired primer-template duplexes was significantly increased for two of the mutants tested. These results suggest that amino acid substitutions that increase the fidelity of dNTP insertion do not necessarily increase misextension fidelity, and that the decreased misextension fidelity may counterbalance the increases in misinsertion fidelity observed for E89G and M184V RTs.

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