Human immunodeficiency virus reverse transcriptase substrate-induced conformational changes and the mechanism of inhibition by nonnucleoside inhibitors.
A combination of transient kinetic and equilibrium titration methods has been used to show that both primer/template and nucleotide binding to human immunodeficiency virus type 1 (HIV-1) reverse transcriptase are two-step processes. In both cases, after initial formation of relatively weakly bound states, isomerization reactions lead to tightly bound states. In the case of deoxynucleotide binding to the reverse transcriptase-primer/template complex, the second step in the interaction is rate-limiting in the overall reaction during processive polymerization. Discrimination against incorrect nucleotides occurs both in the initial weak binding and in the second step but is purely kinetic in the second step (as opposed to thermodynamic in the first step). Nonnucleoside inhibitors have a relatively small effect on nucleotide-binding steps (overall affinity is reduced by a factor of ca. 10), while the affinity of the primer/template duplex is increased by at least a factor of 10. The major effect of nonnucleoside inhibitors is on the chemical step (nucleotide transfer).
Rittinger
K
K
Max-Planck-Institut für Molekulare Physiologie, Abteilung Physikalische Biochemie, Dortmund, Germany.
Divita
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G
Goody
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RS
D000998:Antiviral Agents / Q000494:pharmacology; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology*; D008433:Mathematics; D008962:Models, Theoretical; D008969:Molecular Sequence Data; D011487:Protein Conformation; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry* / Q000378:metabolism*; D018894:Reverse Transcriptase Inhibitors; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D013942:Thymine Nucleotides / Q000378:metabolism
D000998:Antiviral Agents; D017931:DNA Primers; D003845:Deoxycytosine Nucleotides; D004791:Enzyme Inhibitors; D018894:Reverse Transcriptase Inhibitors; D013942:Thymine Nucleotides; C024107:2'-deoxycytidine 5'-triphosphate; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; C024157:thymidine 5'-triphosphate