期刊
IUCRJ
卷 7, 期 -, 页码 976-984出版社
INT UNION CRYSTALLOGRAPHY
DOI: 10.1107/S2052252520012701
关键词
G-protein-coupled receptors; membrane proteins; XFELs; serial femtosecond crystallography; adenosine A(2A) receptors; lipidic cubic phases; high dynamic range detectors
资金
- National Institutes of Health (NIH) [R35 GM127086, R21 DA042298, R01 GM124152]
- National Science Foundation (NSF)
- BioXFEL Science and Technology Center [1231306]
- SLAC National Accelerator Laboratory through the Panofsky fellowship
- Department of Energy through the Panofsky fellowship
- US Department of Energy, Office of Science, Office of Basic Energy Sciences [DE-AC02-76SF00515]
Serial femtosecond crystallography (SFX) with X-ray free-electron lasers (XFELs) has proven highly successful for structure determination of challenging membrane proteins crystallized in lipidic cubic phase; however, like most techniques, it has limitations. Here we attempt to address some of these limitations related to the use of a vacuum chamber and the need for attenuation of the XFEL beam, in order to further improve the efficiency of this method. Using an optimized SFX experimental setup in a helium atmosphere, the room-temperature structure of the adenosine A(2A) receptor (A(2A)AR) at 2.0 angstrom resolution is determined and compared with previous A(2A)AR structures determined in vacuum and/or at cryogenic temperatures. Specifically, the capability of utilizing high XFEL beam transmissions is demonstrated, in conjunction with a high dynamic range detector, to collect high-resolution SFX data while reducing crystalline material consumption and shortening the collection time required for a complete dataset. The experimental setup presented herein can be applied to future SFX applications for protein nanocrystal samples to aid in structure-based discovery efforts of therapeutic targets that are difficult to crystallize.
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