Effects of zidovudine-selected human immunodeficiency virus type 1 reverse transcriptase amino acid substitutions on processive DNA synthesis and viral replication

Angela M. Caliendo, Anu Savara, A. N. Donna, Kimberly DeVore, Joan C. Kaplan, Richard T. D'Aquila*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

89 Scopus citations

Abstract

Certain amino acid substitutions in the reverse transcriptase (RT), including D67N, K70R, T215Y, and K219Q, cause high-level resistance of human immunodeficiency virus type 1 (HIV-1) to zidovudine (3'-azidothymidine; AZT) and appear to approximate the template strand of the enzyme-template-primer complex in structural models. We studied whether this set of mutations altered RT-template-primer interaction as well as their effect on virus replication in the absence of inhibitor. When in vitro polymerization was limited to a single association of an RT with an oligodeoxynucleotide-primed heteropolymeric RNA template (a single processive cycle), recombinant- expressed mutant 67/70/215/219 RT synthesized 5- to 10-fold more high- molecular-weight DNA products (>200 nucleotides in length) than wild-type RT. This advantage was maintained as deoxynucleoside triphosphate (dNTP) concentrations were decreased to limiting levels. In contrast, no difference was seen between wild-type and mutant RTs under conditions allowing repeated associations of enzyme with template-primer. Because intracellular dNTP concentrations are low prior to mitogenic stimulation, we compared replication of mutant 67/70/215/219 virus and wild-type virus in peripheral blood mononuclear cells (PBMC) stimulated before and after infection. In the absence of inhibitor, mutant 67/70/215/219 virus had a replication advantage in PBMC stimulated with phytohemagglutinin and interleukin-2 after infection, but virus replication was similar in PBMC stimulated before infection in vitro. The results confirm that RT mutations D67N, K70R, T215Y, and K219Q affect an enzyme-template-primer interaction in vitro and suggest that such substitutions may affect HIV-1 pathogenesis during therapy by increasing viral replication capacity in cells stimulated after infection.

Original languageEnglish (US)
Pages (from-to)2146-2153
Number of pages8
JournalJournal of virology
Volume70
Issue number4
DOIs
StatePublished - 1996

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

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