Introducing the next revolution
in membrane protein structure determination.

Since the development of lipidic cubic phase (LCP) for the crystallization of membrane proteins in 1996(1), there have been over 200 structures of proteins deposited in the protein data bank using this method(2).  Since its advent, there have been many improvements and advancements to lipidic cubic phase, including robotics, sandwich plates, imaging techniques (e.g. FRAP, SONICC), host lipids, and data collection. Perhaps the next frontier to membrane protein crystallography in lipidic cubic phase is the use of serial femtosecond X-ray crystallography (SFX) to collect data on protein crystals grown with this method (LCP-SFX)(3)
The X-ray free electron laser (XFEL) produces femtosecond X-ray pulses which are nine orders of magnitude brighter than synchrotron sources
(4). Using this instrument, protein microcrystals are introduced to the pulsed XFEL beam via a liquid jet.  When a protein microcrystal comes into contact with the XFEL beam, diffraction of the protein crystal occurs before radiation damage, one of the main advantages of this method.  There are only two XFEL sources able to produce atomic level resolution of protein microcrystals: the Linac Coherent Light Source (LCLS) in Stanford, CA, USA and the SPring-8 Angstrom Compact free electron Laser (SACLA) in Harima, Japan. 

In 2011, using microcrystals of the membrane protein complex photosystem I, the first diffraction images were obtained using SFX(5). Since this breakthrough, many advancements have been made to this technique, including sample delivery, in vivo crystallography, detector technology, and better methods to grow microcrystals. A relatively new development in this field is the ability to deliver protein microcrystals grown in LCP into the XFEL(6). One of the main advantages of LCP-SFX is the ability to use a very low flow rate of the LCP material, which greatly reduces the amount of protein microcrystals needed.

To date, structures of three membrane proteins have been determined with this method: the diacylglycerol kinase DgkA(7), the GPCR serotonin receptor 5-HT2B(8), and the GPCR Human Smoothened Receptor(9). In addition to these membrane protein structures, LCP-SFX technique has also been shown to be applicable to soluble proteins with lysozyme and photocyanin structures being determined with this method(10). Additionally, LCP-SFX technology is being further adapted to perform serial millisecond crystallography on the more widely available synchrotron beamlines(11).


1.       Landau, E. M. and Rosenbusch, J. P. (1996) PNAS 93, 14532-14535.
2.       Caffrey, M. (2015) Acta Cryst F71, 3-18.
3.       Liu, W. et al. (2014) Philos Trans R Soc B 369, 20130314.
4.       Wiener, M. C. (2015) IUCrJ 2, 387-388.
5.       Chapman, H. N. (2011) Nature 470, 73-77.
6.       Liu, W. et al. (2014) Nat Protoc 9, 2123-2134.
7.       Caffrey, M. et al. (2014) Philos Trans R Soc B 369, 20130621.
8.       Liu, W. et al. (2013) Science 342, 1521-1524.
9.       Weierstall, U. et al. (2014) Nat Commun 5, 3309.
10.    Fromme, R. et al. (2015) IUCrJ 2, 545-551.
11.    Nogly, P. et al. (2015), IUCrJ 2, 168-176.