Conformational Plasticity of the NNRTI-Binding Pocket in HIV-1 Reverse Transcriptase: A Fluorine Nuclear Magnetic Resonance Study

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• 作者：Naima G. Sharaf ; Rieko Ishima ; Angela M. Gronenborn
• 刊名：Biochemistry
• 出版年：2016
• 出版时间：July 19, 2016
• 年：2016
• 卷：55
• 期：28
• 页码：3864-3873
• 全文大小：589K
• 年卷期：0
• ISSN：1520-4995

文摘

HIV-1 reverse transcriptase (RT) is a major drug target in the treatment of HIV-1 infection. RT inhibitors currently in use include non-nucleoside, allosteric RT inhibitors (NNRTIs), which bind to a hydrophobic pocket, distinct from the enzyme’s active site. We investigated RT–NNRTI interactions by solution ¹⁹F nuclear magnetic resonance (NMR), using singly ¹⁹F-labeled RT proteins. Comparison of ¹⁹F chemical shifts of fluorinated RT and drug-resistant variants revealed that the fluorine resonance is a sensitive probe for identifying mutation-induced changes in the enzyme. Our data show that in the unliganded enzyme, the NNRTI-binding pocket is highly plastic and not locked into a single conformation. Upon inhibitor binding, the binding pocket becomes rigidified. In the inhibitor-bound state, the ¹⁹F signal of RT is similar to that of drug-resistant mutant enzymes, distinct from what is observed for the free state. Our results demonstrate the power of ¹⁹F NMR spectroscopy to characterize conformational properties using selectively ¹⁹F-labeled protein.