title: Application of Site-Specific Spin Labeling for NMR Detecting Inhibitor-Induced Conformational Change of HIV-1 Reverse Transcriptase.
abstract: Paramagnetism-assisted nuclear magnetic resonance (NMR) techniques can provide long-range structural information complemented with local information derived from chemical-shift perturbation and nuclear Overhauser effect data. Here, we address the application of paramagnetic relaxation enhancement (PRE) to detect inhibitor-induced conformational change of a drug target protein using human immunodeficiency virus type 1 reverse transcriptase (HIV-1 RT) as a model protein. Using a site-specific spin-labeled HIV-1 RT mutant with selective (13) C labeling, conformation-dependent PREs were successfully observed reflecting the stabilization of an open conformation of this enzyme caused by inhibitor binding. This study demonstrates that the paramagnetism-assisted NMR approach offers an alternative strategy in protein-based drug screening to identify allosteric inhibitors of a target protein.
authors.affiliation: Graduate Program in Bioscience, Faculty of Science, Graduate School, Kasetsart University, Chatuchak, Bangkok, 10900, Thailand.|Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.
authors.forename: Supaporn
authors.initials: S
authors.lastname: Seetaha
authors.affiliation: Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.|Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.
authors.forename: Maho
authors.initials: M
authors.lastname: Yagi-Utsumi
authors.affiliation: Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.|Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.|School of Materials Science, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, 923-1292, Japan.
authors.forename: Takumi
authors.initials: T
authors.lastname: Yamaguchi
authors.affiliation: Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Ishii
authors.affiliation: Department of Chemistry, Faculty of Science, Kasetsart University, Chatuchak, Bangkok, 10900, Thailand.
authors.forename: Supa
authors.initials: S
authors.lastname: Hannongbua
authors.affiliation: Department of Biochemistry, Faculty of Science, Kasetsart University, Chatuchak, Bangkok, 10900, Thailand. fsciktc@ku.ac.th.|Center for Advanced Studies in Tropical Natural Resources, National Research University-Kasetsart University, Chatuchak, Bangkok, 10900, Thailand. fsciktc@ku.ac.th.
authors.forename: Kiattawee
authors.initials: K
authors.lastname: Choowongkomon
authors.affiliation: Institute for Molecular Science, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan. kkato@phar.nagoya-cu.ac.jp.|Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, 5-1 Higashiyama, Mhodaiji, Okazaki, 444-8787, Japan. kkato@phar.nagoya-cu.ac.jp.|Graduate School of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabe-dori, Mizuhoku, Nagoya, 467-8603, Japan. kkato@phar.nagoya-cu.ac.jp.
authors.forename: Koichi
authors.initials: K
authors.lastname: Kato
mesh_terms: D002250:Carbon Radioisotopes / Q000032:analysis; D055808:Drug Discovery; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D015497:HIV-1 / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation / Q000187:drug effects*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*
chemical_list: D002250:Carbon Radioisotopes; D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase
keywords: HIV-1 reverse transcriptase; NMR spectroscopy; drug discovery; high-throughput screening; spin labeling