期刊
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
卷 18, 期 9, 页码 -出版社
MDPI
DOI: 10.3390/ijms18091918
关键词
XFEL; SFX; rsFP; Skylan-NS; SACLA; LCLS
资金
- Engineering and Physical Sciences Research Council (EPSRC) [EP/M000192/1]
- Biotechnology and Biological Sciences Research Council (BBSRC) [BB/P00752X/1]
- Engineering and Physical Sciences Research Council (UK) (EPSRC) [EP/I032517/1]
- European Research Council (ERC) ASTEX Project [290467]
- Science and Technology Facilities Council (STFC)
- Marie Curie ITN [EC317232]
- U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences [DE-AC02-76SF00515]
- Biotechnology and Biological Sciences Research Council [BB/P00752X/1] Funding Source: researchfish
- Engineering and Physical Sciences Research Council [EP/I032517/1, EP/M000192/1] Funding Source: researchfish
- BBSRC [BB/P00752X/1] Funding Source: UKRI
- EPSRC [EP/I032517/1, EP/M000192/1] Funding Source: UKRI
The photochromic fluorescent protein Skylan-NS (Nonlinear Structured illumination variant mEos3.1H62L) is a reversibly photoswitchable fluorescent protein which has an unilluminated/ground state with an anionic and cis chromophore conformation and high fluorescence quantum yield. Photo-conversion with illumination at 515 nm generates a meta-stable intermediate with neutral trans-chromophore structure that has a 4 h lifetime. We present X-ray crystal structures of the cis (on) state at 1.9 Angstrom resolution and the trans (off) state at a limiting resolution of 1.55 Angstrom from serial femtosecond crystallography experiments conducted at SPring-8 Angstrom Compact Free Electron Laser (SACLA) at 7.0 keV and 10.5 keV, and at Linac Coherent Light Source (LCLS) at 9.5 keV. We present a comparison of the data reduction and structure determination statistics for the two facilities which differ in flux, beam characteristics and detector technologies. Furthermore, a comparison of droplet on demand, grease injection and Gas Dynamic Virtual Nozzle (GDVN) injection shows no significant differences in limiting resolution. The photoconversion of the on- to the off-state includes both internal and surface exposed protein structural changes, occurring in regions that lack crystal contacts in the orthorhombic crystal form.
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