NDLI: Mechanism of allosteric inhibition of HIV-1 reverse transcriptase revealed by single-molecule and ensemble fluorescence

Content Provider	PubMed Central
Author	Schauer, Grant D. Huber, Kelly D. Leuba, Sanford H. Nicolas, Sluis-cremer
Copyright Year	2014
Abstract	Non-nucleoside reverse transcriptase (RT) inhibitors (NNRTIs) are routinely used to treat HIV-1 infection, yet their mechanism of action remains unclear despite intensive investigation. In this study, we developed complementary single-molecule fluorescence and ensemble fluorescence anisotropy approaches to discover how NNRTIs modulate the intra-molecular conformational changes and inter-molecular dynamics of RT-template/primer (T/P) and RT–T/P–dNTP complexes. We found that NNRTI binding to RT induces opening of the fingers and thumb subdomains, which increases the dynamic sliding motion of the enzyme on the T/P and reduces dNTP binding affinity. Further, efavirenz promotes formation of the E138-K101 salt bridge between the p51 and p66 subunits of RT, which contributes to opening of the thumb/fingers subdomains. Engineering a more polar salt bridge between p51 and p66 resulted in even greater increases in the thumb/fingers opening, RT sliding, dNTP binding disruption and in vitro and in vivo RT inhibition than were observed with wild-type RT. We also observed that K103N, a clinically relevant NNRTI resistance mutation, does not prevent binding between efavirenz and RT-T/P but instead allows formation of a stable and productive RT–T/P–dNTP complex, possibly through disruption of the E138-K101 salt bridge. Collectively, these data describe unique structure–activity–resistance relationships that could be exploited for drug development.
Related Links	http://dx.doi.org/10.1093/nar/gku819
Ending Page	11696
Page Count	10
Starting Page	11687
File Format	PDF
ISSN	03051048
e-ISSN	13624962
Journal	Nucleic Acids Research
Issue Number	18
Volume Number	42
Language	English
Publisher	Oxford University Press
Publisher Date	2014-10-13
Access Restriction	Open
Rights Holder	Oxford University Press
Subject Keyword	Genetics Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Genetics

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