Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

Nakane Takanori; Hanashima Shinya; Suzuki MamoruSaiki HarukaHayashi TaichiKakinouchi KeisukeSugiyama ShigeruKawatake SatoshiMatsuoka ShigeruMatsumori NobuakiNango ErikoKobayashi JunShimamura TatsuroKimura KanakoMori ChihiroKunishima NaokiSugahara MichihiroTakakyu YokoInoue ShigeyukiMasuda TetsuyaHosaka ToshiakiTono KensukeJoti YasumasaKameshima TakashiHatsui TakakiYabashi MakinaInoue TsuyoshiNureki OsamuIwata SoMurata MichioMizohata Eiichi

首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America. >Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

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Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

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The 3D structure determination of biological macromolecules by X-ray crystallography suffers from a phase problem: to perform Fourier transformation to calculate real space density maps, both intensities and phases of structure factors are necessary; however, measured diffraction patterns give only intensities. Although serial femtosecond crystallography (SFX) using X-ray free electron lasers (XFELs) has been steadily developed since 2009, experimental phasing still remains challenging. Here, using 7.0-keV (1.771 angstrom)X-ray pulses from the SPring-8 Angstrom Compact Free Electron Laser (SACLA), iodine single-wavelength anomalous diffraction (SAD), single isomorphous replacement (SIR), and single isomorphous replacement with anomalous scattering (SIRAS) phasing were performed in an SFX regime for a model membrane protein bacteriorhodopsin (bR). The crystals grown in bicelles were derivatized with an iodine-labeled detergent heavy-atom additive 13a (HAD13a), which contains the magic triangle, I3C head group with three iodine atoms. The alkyl tail was essential for binding of the detergent to the surface of bR. Strong anomalous and isomorphous difference signals from HAD13a enabled successful phasing using reflections up to 2.1- resolution from on 3,000 and 4,000 indexed images from native and derivative crystals, respectively. When more images were merged, structure solution was possible with data truncated at 3.3-angstrom resoluti, which is the lowest resolution among the reported cases of SFX phasing. Moreover, preliminary SFX experiment showed that HAD13a successfully derivatized the G protein-coupled A2a adenosine receptor crystallized in lipidic cubic phases. These results pave the way for de novo structure determination of membrane proteins, which often diffract poorly, even with the brightest XFEL beams.

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《Proceedings of the National Academy of Sciences of the United States of America.》 |2016年第46期|13039-13044|共6页
作者
Nakane Takanori; Hanashima Shinya; Suzuki MamoruSaiki HarukaHayashi TaichiKakinouchi KeisukeSugiyama ShigeruKawatake SatoshiMatsuoka ShigeruMatsumori NobuakiNango ErikoKobayashi JunShimamura TatsuroKimura KanakoMori ChihiroKunishima NaokiSugahara MichihiroTakakyu YokoInoue ShigeyukiMasuda TetsuyaHosaka ToshiakiTono KensukeJoti YasumasaKameshima TakashiHatsui TakakiYabashi MakinaInoue TsuyoshiNureki OsamuIwata SoMurata MichioMizohata Eiichi;
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作者单位

Univ Tokyo, Grad Sch Sci, Dept Biol Sci, Tokyo 1130033, Japan;

Osaka Univ, Grad Sch Sci, Dept Chem, Osaka 5630043, Japan;

Osaka Univ, Inst Prot Res, Osaka 5650871, JapanOsaka Univ, Grad Sch Engn, Dept Appl Chem, Osaka 5650871, JapanRIKEN, SPring Ctr 8, Sayo, Hyogo 6795148, JapanKyoto Univ, Grad Sch Med, Dept Cell Biol, Kyoto 6068501, JapanRIKEN, Ctr Life Sci Technol, Div Struct & Synthet Biol, Yokohama, Kanagawa 2300045, JapanJapan Synchrotron Radiat Res Inst, Sayo, Hyogo 6795198, Japan;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
phase problem; de novo phasing; heavy atom; detergent; G protein-coupled receptor;

Membrane protein structure determination by SAD, SIR, or SIRAS phasing in serial femtosecond crystallography using an iododetergent

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