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SEMINAR
Solid-state NMR study of proteins and the signal enhancement by DNP using unpaired electron spins
Abstract
NMR sensitivity is the crucial factor for studying the structures of large biomolecules. We are improving the instrumentation for high-field dynamic nuclear polarization (DNP) to increase the sensitivity of high-resolution solid-state 13C NMR. We will present the instrumentation for the DNP-NMR spectrometer for 700 MHz proton and 460 GHz electron resonances under magic-angle spinning at 30 K, and the experimental results. Magic-angle-spinning solid-state 13C NMR spectroscopy is useful for structural analysis of non-crystalline proteins. However, the signal assignments and structural analysis are often hampered by the signal overlaps primarily due to minor structural heterogeneities, especially for uniformly-13C,15N labeled samples at low temperatures for DNP. To overcome this problem, we presented a method for assigning 13C chemical shifts and secondary structures from unresolved two-dimensional 13C−13C MAS NMR spectra by spectral fitting.
References
Matsuki, Y., et al. (2012) Helium-cooling and -spinning dynamic nuclear polarization for sensitivity-enhanced solid-state NMR at 14 T and 30 K. J. Magn. Reson., 225, 1-9.
Ikeda, K., et al. (2011) Combined use of replica-exchange molecular dynamics and magic-angle-spinning solid-state NMR spectral simulations for determining the structure and orientation of membrane-bound peptide. J. Phys. Chem. B, 115, 9327-9336.
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BIOGRAPHY
2008-present: Professor, Professor at Institute for Protein Research, Osaka Univ.
2001-2008: Associate Professor, Professor at Institute for Protein Research, Osaka Univ.
1992-2000: Lecturer and Associate Professor, Department of Chemistry and Biotechnology, Yokohama National Univ.
1985-1992: Scientist, Biological Metrology Lab, JEOL Ltd.
1985: Dr. Sci., Chemistry, Graduate School of Science, Osaka Univ.
Research Fields and Interests:
NMR provides information on the protein structure at work with atomic resolution even in cells. Taking this advantage, we are studying the biological activities for signal transduction and energy conversion from structures. Since supramolecular systems such as membrane protein complexes play important roles in biological systems, we are also developing new methodologies in NMR to analyze those challenging structures. One our method for solid-state NMR features dynamic nuclear polarization, DNP, for a 1000-fold signal enhancement using high-intensity submillimeter wave gyrotrons. We aim to contribute to academic and industrial NMR applications such as drug discovery.
Selected Publications
Matsuki, Y., Nakamura, S., Fukui, S., Suematsu, H., Fujiwara T. (2015) Closed-cycle cold helium magic-angle spinning for sensitivity-enhanced multi-dimensional solid-state NMR. J. Magn. Reson. in press.
Furuita, K., Kataoka, S., Sugiki, T., Hattori, Y., Kobayashi, N., Ikegami, T., Shiozaki, K., Fujiwara, T., Kojima, C. (2015) Utilization of paramagnetic relaxation enhancements for high-resolution NMR structure determination of a soluble loop-rich protein with sparse NOE distance restraints. J. Biomol. NMR, 61, 55-64.
Kang, S.-J., Todokoro, Y., Yumen, I., Shen, B., Iwasaki, I., Suzuki, T., Miyagi, A., Yoshida, M., Fujiwara, T., Akutsu, H. (2014) Active-site structure of thermophilic Foc subunit ring in membranes elucidated by solid-state NMR, Biophysical J., 106, 390-398.
Ikeda, K., Egawa, A., Fujiwara, T. (2013) Secondary structural analysis of proteins based on 13C chemical shift assignments in unresolved solid-state NMR spectra Enhanced by Fragmented Structure Database. J. Biomol. NMR, 55, 189-200.
Matsuki, Y., Ueda, K., Idehara, T., Ikeda, R., Kosuga, K., Ogawa, I., Nakamura, S., Toda, M., Anai, T., Fujiwara, T. (2012) Application of continuously frequency-tunable 0.4 THz gyrotron to dynamic nuclear polarization for 600 MHz solid-state NMR. J. Infrared Millim. Terahz. Waves, 33, 745-755.
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