TY - JOUR
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.
N1 - Funding Information:
The authors wish to thank John Blanchard and Lisa Rezende for reading the manuscript and the Oligonucleotide Synthesis Facility of the Albert Einstein College of Medicine’s Cancer Center for DNA oligonucleotides. Data in this paper are from a thesis to be submitted in partial fulfillment of the requirements for the Degree of Doctor of Philosophy in the Sue Golding Graduate Division of Medical Sciences, Albert Einstein College of Medicine, Yeshiva University. This work was supported by Public Health Service grants AI30861 and AI40375 (to V.R.P.). M.E.H. and W.C.D. would like to acknowledge support from the institutional training grants NIGMS T32-GM07491 and NIAID T32-AI07501, respectively.
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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U2 - 10.1093/nar/26.19.4389
DO - 10.1093/nar/26.19.4389
M3 - Article
C2 - 9742239
AN - SCOPUS:0032189086
SN - 0305-1048
VL - 26
SP - 4389
EP - 4394
JO - Nucleic acids research
JF - Nucleic acids research
IS - 19
ER -