Preparation of microcrystals in lipidic cubic phase for serial femtosecond crystallography

Wei Liu, Andrii Ishchenko, Vadim Cherezov

Research output: Contribution to journalArticle

71 Scopus citations

Abstract

We have recently established a procedure for serial femtosecond crystallography (SFX) in lipidic cubic phase (LCP) for protein structure determination at X-ray free-electron lasers (XFELs). LCP-SFX uses the gel-like LCP as a matrix for growth and delivery of membrane protein microcrystals for crystallographic data collection. LCP is a liquid-crystalline mesophase composed of lipids and water. It provides a membrane-mimicking environment that stabilizes membrane proteins and supports their crystallization. Here we describe detailed procedures for the preparation and characterization of microcrystals for LCP-SFX applications. The advantages of LCP-SFX over traditional crystallographic methods include the capability of collecting room-temperature high-resolution data with minimal effects of radiation damage from sub-10-μm crystals of membrane and soluble proteins that are difficult to crystallize, while eliminating the need for crystal harvesting and cryo-cooling. Compared with SFX methods for microcrystals in solution using liquid injectors, LCP-SFX reduces protein consumption by 2-3 orders of magnitude for data collection at currently available XFELs. The whole procedure typically takes 3-5 d, including the time required for the crystals to grow.

Original languageEnglish (US)
Pages (from-to)2123-2134
Number of pages12
JournalNature Protocols
Volume9
Issue number9
DOIs
StatePublished - 2014
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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