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Structure determination of a novel protein by sulphur SAD using chromium radiation in combination with a new crystal mounting method

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Title: Structure determination of a novel protein by sulphur SAD using chromium radiation in combination with a new crystal mounting method
Authors: Kitago, Yu Browse this author
Watanabe, Nobuhisa Browse this author
Tanaka, Isao Browse this author →KAKEN DB
Keywords: sulphur
single-wavelength anomalous scattering
chromium radiation
crystal mounting method
Issue Date: Aug-2005
Publisher: International Union of Crystallography
Journal Title: Acta Crystallographica Section D
Volume: 61
Issue: 8
Start Page: 1013
End Page: 1021
Publisher DOI: 10.1107/S0907444905012734
PMID: 16041065
Abstract: A novel and easy crystal mounting technique was developed for the sulphur SAD method using chromium Kα radiation (2.29Å). Using this technique, the cryo-buffer and cryoloop around the protein crystal can be removed before data collection to eliminate their X-ray absorption. The superiority and the reproducibility of the datasets with our mounting technique were demonstrated using tetragonal hen egg-white lysozyme crystals. The structure of a novel protein, PH1109, from Pyrococcus horikoshii OT3 was solved using this technique. At the wavelength of chromium Kα radiation, anomalous signals, <|ΔF|>/<|F|> of PH1109 is expected to be 1.72%, as this protein of 144 residues includes 4 methionines and 2 cysteines. Sulphur SAD phasing was performed using SHELXD and SHELXE. In the case of the dataset obtained using our novel crystal mounting technique, 54.9 % of all residues were built with side-chains automatically by RESOLVE. On the other hand, only 16.0 % were built with side-chains for the dataset collected using the standard cryoloop. These results indicated that our crystal mounting technique was superior to the standard loop mounting method for measurement of small anomalous differences at longer wavelength, and yielded better results in sulphur substructure solution and initial phasing. The present study demonstrated that the sulphur SAD method with a chromium source becomes enhanced more practical for macromolecular structure determination using our crystal mounting technique.
Rights: Copyright © International Union of Crystallography
Type: article (author version)
Appears in Collections:生命科学院・先端生命科学研究院 (Graduate School of Life Science / Faculty of Advanced Life Science) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)

Submitter: 渡邉 信久

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