An isomorphous replacement method for efficient de novo phasing for serial femtosecond crystallography

• Published in: Scientific reports Vol. 5; no. 1; p. 14017
• Main Authors: Yamashita, Keitaro, Pan, Dongqing, Okuda, Tomohiko, Sugahara, Michihiro, Kodan, Atsushi, Yamaguchi, Tomohiro, Murai, Tomohiro, Gomi, Keiko, Kajiyama, Naoki, Mizohata, Eiichi, Suzuki, Mamoru, Nango, Eriko, Tono, Kensuke, Joti, Yasumasa, Kameshima, Takashi, Park, Jaehyun, Song, Changyong, Hatsui, Takaki, Yabashi, Makina, Iwata, So, Kato, Hiroaki, Ago, Hideo, Yamamoto, Masaki, Nakatsu, Toru
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 11-09-2015; Nature Publishing Group
• Subjects: 631/535/1266; 639/624/1020/1087; Crystallography; Crystallography, X-Ray; Crystals; Data collection; Data processing; Humanities and Social Sciences; Lasers; Models, Molecular; multidisciplinary; Proteins - chemistry; Science
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep14017

Serial femtosecond crystallography (SFX) with X-ray free electron lasers (XFELs) holds great potential for structure determination of challenging proteins that are not amenable to producing large well diffracting crystals. Efficient
de novo
phasing methods are highly demanding and as such most SFX structures have been determined by molecular replacement methods. Here we employed single isomorphous replacement with anomalous scattering (SIRAS) for phasing and demonstrate successful application to SFX
de novo
phasing. Only about 20,000 patterns in total were needed for SIRAS phasing while single wavelength anomalous dispersion (SAD) phasing was unsuccessful with more than 80,000 patterns of derivative crystals. We employed high energy X-rays from SACLA (12.6 keV) to take advantage of the large anomalous enhancement near the
L
III
absorption edge of Hg, which is one of the most widely used heavy atoms for phasing in conventional protein crystallography. Hard XFEL is of benefit for
de novo
phasing in the use of routinely used heavy atoms and high resolution data collection.