Protonation/Deprotonation of Proteins by Neutron Diffraction Structure Analysis
Neutron protein crystallography can reveal nuclear position and it is very useful to find hydrogen or protonation/deprotonation of protein. It is, however, an intensity-limited experiment and requires large and good quality single protein crystal, so the user population has been so small. Recently, new intense neutron source makes ones to find several protonation states in proteins; PcyA complex (complex of Phycocyanobilin: Ferredoxin Oxidoreductase and Biliverdin IXα), cellulase and substrate complex and farnesyl pyrophosphate synthase (FPPS)-drug complex. At the same time, new techniques for neutron measurement such as high pressure freezing and dynamic nuclear polarization of protein have been also tried to be developed. Finally, a plan of new neutron facility to gain more S/N ratio is expected so that the sample crystal volume can be much small to find protonation/deprotonation.
KeywordsNeutron protein crystallography Proton Dynamic nuclear polarization High-pressure freezing Spallation neutron source
Authors would thank Profs. T. Iwata and Miyachi and their students in Yamagata University for conducting and discussing ESR and DNP measurements, Prof. Chatake in Kyoto University for preparation of protein crystals and discussion, Dr. T. Kumada in Japan Atomic Energy Agency (JAEA) for providing a radical concentration information, and Drs. H. Seto in High Energy Accelerator Research Organization (KEK) and Harada in JAEA for providing information of the second target station at MLF in J-PARC. Synchrotron radiation experiment was conducted at BL5A of the Photon Factory in KEK, Ibaraki, Japan [2014G650]. Finally, authors are profoundly grateful to iBIX users, Prof. T. Yamada, Dr. N. Yano, Mrs. S. Ninomiya and J. Hiroki, and all students in Tanaka laboratory in Ibaraki University.
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