The K65R mutant reverse transcriptase of HIV-1 cross-resistant to 2', 3'-dideoxycytidine, 2',3'-dideoxy-3'-thiacytidine, and 2',3'-dideoxyinosine shows reduced sensitivity to specific dideoxynucleoside triphosphate inhibitors in vitro

J Biol Chem. 1994 Nov 11;269(45):28118-22.


The K65R mutation in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) encodes cross-resistance to 2',3'-dideoxycytidine (ddC), 2',3'-dideoxy-3'-thiacytidine (3TC), and 2',3'-dideoxyinosine (ddI). We characterized the in vitro sensitivities of recombinant wild type (wt) and K65R mutant RT to dideoxynucleoside triphosphate (ddNTP) inhibitors, using a variety of primer-templates. With poly(rA)-oligo(dT), the K65R mutant showed slight increases in Ki for ddTTP and 3'-azido, 3'-deoxythymidine triphosphate (AZTTP) compared to wt RT, but neither wt nor K65R RT was inhibited by ddCTP or ddATP. With poly(rI)-oligo(dC), the K65R mutant showed a 2-fold increase in Km for dCTP and a 20-fold increase in Ki for ddCTP compared to wt, whereas ddATP, ddTTP, and AZTTP failed to inhibit either enzyme. With a heteropolymeric primer-template, the K65R mutant showed 10-fold reduced sensitivities to ddCTP, 3TCTP, and ddATP, and 4-fold reduced sensitivity to AZTTP, compared to wt. In contrast, both enzymes were equally inhibited by ddTTP and ddGTP. HIV-1 cross-resistance to ddC/3TC/ddI resulting from the K65R mutation may therefore involve selective alterations in substrate/inhibitor recognition. Additionally, competitive inhibition by ddNTPs noncomplementary to the template base appears to be unimportant in the mechanism of inhibition of HIV-1 RT by dideoxynucleoside analogs.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antiviral Agents / pharmacology*
  • Base Sequence
  • Cloning, Molecular
  • DNA Primers
  • Didanosine / pharmacology*
  • Drug Resistance, Microbial
  • HIV Reverse Transcriptase
  • HIV-1 / drug effects
  • HIV-1 / enzymology*
  • Kinetics
  • Lamivudine
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Point Mutation*
  • Polynucleotides
  • RNA-Directed DNA Polymerase / isolation & purification
  • RNA-Directed DNA Polymerase / metabolism*
  • Recombinant Proteins / antagonists & inhibitors
  • Recombinant Proteins / metabolism
  • Reverse Transcriptase Inhibitors
  • Substrate Specificity
  • Templates, Genetic
  • Zalcitabine / analogs & derivatives*
  • Zalcitabine / pharmacology


  • Antiviral Agents
  • DNA Primers
  • Polynucleotides
  • Recombinant Proteins
  • Reverse Transcriptase Inhibitors
  • Lamivudine
  • Zalcitabine
  • HIV Reverse Transcriptase
  • RNA-Directed DNA Polymerase
  • Didanosine