title: Development of a dose-limiting data collection strategy for serial synchrotron rotation crystallography.
abstract: Serial crystallography, in which single-shot diffraction images are collected, has great potential for protein microcrystallography. Although serial femtosecond crystallography (SFX) has been successfully demonstrated, limited beam time prevents its routine use. Inspired by SFX, serial synchrotron crystallography (SSX) has been investigated at synchrotron macromolecular crystallography beamlines. Unlike SFX, the longer exposure time of milliseconds to seconds commonly used in SSX causes radiation damage. However, in SSX, crystals can be rotated during the exposure, which can achieve efficient coverage of the reciprocal space. In this study, mercury single-wavelength anomalous diffraction (Hg-SAD) phasing of the luciferin regenerating enzyme (LRE) was performed using serial synchrotron rotation crystallography. The advantages of rotation and influence of dose on the data collected were evaluated. The results showed that sample rotation was effective for accurate data collection, and the optimum helical rotation step depended on multiple factors such as multiplicity and partiality of reflections, exposure time per rotation angle and the contribution from background scattering. For the LRE microcrystals, 0.25° was the best rotation step for the achievable resolution limit, whereas a rotation step larger than or equal to 1° was favorable for Hg-SAD phasing. Although an accumulated dose beyond 1.1 MGy caused specific damage at the Hg site, increases in resolution and anomalous signal were observed up to 3.4 MGy because of a higher signal-to-noise ratio.
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo 679-5198, Japan.
authors.forename: Kazuya
authors.initials: K
authors.lastname: Hasegawa
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Keitaro
authors.initials: K
authors.lastname: Yamashita
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo, Kyoto 606-8501, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Murai
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo 679-5198, Japan.
authors.forename: Nipawan
authors.initials: N
authors.lastname: Nuemket
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Kunio
authors.initials: K
authors.lastname: Hirata
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Go
authors.initials: G
authors.lastname: Ueno
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Hideo
authors.initials: H
authors.lastname: Ago
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Nakatsu
authors.affiliation: Japan Synchrotron Radiation Research Institute, 1-1-1 Kouto, Sayo 679-5198, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kumasaka
authors.affiliation: RIKEN SPring-8 Center, 1-1-1 Kouto, Sayo 679-5148, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Yamamoto
keywords: mercury single-wavelength anomalous diffraction; radiation damage; serial synchrotron crystallography