Post-Catalytic Complexes with Emtricitabine or Stavudine and HIV-1 Reverse Transcriptase Reveal New Mechanistic Insights for Nucleotide Incorporation and Drug Resistance.

Nicole Bertoletti,Karen S Anderson,Raymond F Schinazi,Albert H Chan

doi:10.3390/molecules25204868

Abstract

Human immunodeficiency virus 1 (HIV-1) infection is a global health issue since neither a cure nor a vaccine is available. However, the highly active antiretroviral therapy (HAART) has improved the life expectancy for patients with acquired immunodeficiency syndrome (AIDS). Nucleoside reverse transcriptase inhibitors (NRTIs) are in almost all HAART and target reverse transcriptase (RT), an essential enzyme for the virus. Even though NRTIs are highly effective, they have limitations caused by RT resistance. The main mechanisms of RT resistance to NRTIs are discrimination and excision. Understanding the molecular mechanisms for discrimination and excision are essential to develop more potent and selective NRTIs. Using protein X-ray crystallography, we determined the first crystal structure of RT in its post-catalytic state in complex with emtricitabine, (-)FTC or stavudine (d4T). Our structural studies provide the framework for understanding how RT discriminates between NRTIs and natural nucleotides, and for understanding the requirement of (-)FTC to undergo a conformation change for successful incorporation by RT. The crystal structure of RT in post-catalytic complex with d4T provides a “snapshot” for considering the possible mechanism of how RT develops resistance for d4T via excision. The findings reported herein will contribute to the development of next generation NRTIs.

Highlights

Since its discovery in the early 1980s, the human immunodeficiency virus 1 (HIV-1) has been a major health issue with nearly 38.0 million people infected globally in 2019 according to the WHO [1]
A crystal structure of reverse transcriptase (RT) in complex with a DNA primer not chain terminated with deoxyguanosine triphosphate in the priming site (P site), and d4T monophosphate (d4T-MP) or (-)FTC monophosphate
A water molecule is bridging an interaction between the α-phosphate group of the d4T-MP incorporated in the DNA primer and the α-phosphate group of d4T-TP in the nucleotide-binding site (N site)

Summary

Introduction

Since its discovery in the early 1980s, the human immunodeficiency virus 1 (HIV-1) has been a major health issue with nearly 38.0 million people infected globally in 2019 according to the WHO [1]. Two of the six classes of the United States Food and Drug Administration (FDA) approved drugs for HIV-1 treatment target the reverse transcriptase (RT) protein, an enzyme critical for the replication cycle of HIV-1 [2]. These two classes are nucleoside RT inhibitors (NRTIs) and non-nucleoside RT. While NNRTIs are allosteric inhibitors that alter the chemical catalysis rate limiting step through conformational changes [3,4], NRTIs mimic nucleotides that bind to the active site of RT. NRTIs are essential components of HAART and part of almost all FDA approved combination therapies for the treatment and protection of an infection with HIV. Some FDA-approved NRTIs are rarely prescribed (e.g., stavudine (d4T)) or discontinued (e.g., zalcitabine (ddC)) due to their off-target toxicity [6]

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Conclusion