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      Solid-State NMR Provides Evidence for Small-Amplitude Slow Domain Motions in a Multispanning Transmembrane α-Helical Protein

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          Abstract

          Proteins are dynamic entities and populate ensembles of conformations. Transitions between states within a conformational ensemble occur over a broad spectrum of amplitude and time scales, and are often related to biological function. Whereas solid-state NMR (SSNMR) spectroscopy has recently been used to characterize conformational ensembles of proteins in the microcrystalline states, its applications to membrane proteins remain limited. Here we use SSNMR to study conformational dynamics of a seven-helical transmembrane (TM) protein, Anabaena Sensory Rhodopsin (ASR) reconstituted in lipids. We report on site-specific measurements of the 15N longitudinal R 1 and rotating frame R relaxation rates at two fields of 600 and 800 MHz and at two temperatures of 7 and 30 °C. Quantitative analysis of the R 1 and R values and of their field and temperature dependencies provides evidence of motions on at least two time scales. We modeled these motions as fast local motions and slower collective motions of TM helices and of structured loops, and used the simple model-free and extended model-free analyses to fit the data and estimate the amplitudes, time scales and activation energies. Faster picosecond (tens to hundreds of picoseconds) local motions occur throughout the protein and are dominant in the middle portions of the TM helices. In contrast, the amplitudes of the slower collective motions occurring on the nanosecond (tens to hundreds of nanoseconds) time scales, are smaller in the central parts of helices, but increase toward their cytoplasmic sides as well as in the interhelical loops. ASR interacts with a soluble transducer protein on its cytoplasmic surface, and its binding affinity is modulated by light. The larger amplitude of motions on the cytoplasmic side of the TM helices correlates with the ability of ASR to undergo large conformational changes in the process of binding/unbinding the transducer.

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          Model-free approach to the interpretation of nuclear magnetic resonance relaxation in macromolecules. 1. Theory and range of validity

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            An improved broadband decoupling sequence for liquid crystals and solids.

            Recently we developed an efficient broadband decoupling sequence called SPARC-16 for liquid crystals ¿J. Magn. Reson. 130, 317 (1998). The sequence is based upon a 16-step phase cycling of the 2-step TPPM decoupling method for solids ¿J. Chem. Phys. 103, 6951 (1995). Since then, we have found that a stepwise variation of the phase angle in the TPPM sequence offers even better results. The application of this new method to a liquid crystalline compound, 4-n-pentyl-4'-cyanobiphenyl, and a solid, L-tyrosine hydrochloride, is reported. The reason for the improvement is explained by an analysis of the problem in the rotating frame. Copyright 2000 Academic Press.
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              Proton-enhanced NMR of dilute spins in solids

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                Author and article information

                Journal
                J Am Chem Soc
                J. Am. Chem. Soc
                ja
                jacsat
                Journal of the American Chemical Society
                American Chemical Society
                0002-7863
                1520-5126
                14 June 2017
                12 July 2017
                : 139
                : 27
                : 9246-9258
                Affiliations
                [1] Department of Physics and Biophysics Interdepartmental Group, University of Guelph , Guelph, Ontario N1G 2W1, Canada
                [§ ]Department of Chemistry, University of Warwick , Coventry CV4 7AL, United Kingdom
                Author notes
                Article
                10.1021/jacs.7b03974
                5510093
                28613900
                163f0a1a-0c44-4015-8fe0-4f459733e2e4
                Copyright © 2017 American Chemical Society

                This is an open access article published under a Creative Commons Attribution (CC-BY) License, which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited.

                History
                : 19 April 2017
                Categories
                Article
                Custom metadata
                ja7b03974
                ja-2017-03974b

                Chemistry
                Chemistry

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