Characterization of lipid matrices for membrane protein crystallization by high-throughput small angle X-ray scattering

Jeremiah S. Joseph, Wei Liu, Joshua Kunken, Thomas M. Weiss, Hiro Tsuruta, Vadim Cherezov

Research output: Contribution to journalArticle

26 Scopus citations

Abstract

The lipidic cubic phase (LCP) has repeatedly proven to serve as a successful membrane-mimetic matrix for a variety of difficult-to-crystallize membrane proteins. While monoolein has been the predominant lipid of choice, there is a growing need for the characterization and use of other LCP host lipids, allowing exploration of a range of structural parameters such as bilayer thickness and curvature for optimal insertion, stability and crystallogenesis of membrane proteins. Here, we describe the development of a high-throughput (HT) pipeline to employ small angle X-ray scattering (SAXS) - the most direct technique to identify lipid mesophases and measure their structural parameters - to interrogate rapidly a large number of lipid samples under a variety of conditions, similar to those encountered during crystallization. Leveraging the identical setup format for LCP crystallization trials, this method allows the quickly assessment of lipid matrices for their utility in membrane protein crystallization, and could inform the tailoring of lipid and precipitant conditions to overcome specific crystallization challenges. As proof of concept, we present HT LCP-SAXS analysis of lipid samples made of monoolein with and without cholesterol, and of monovaccenin, equilibrated with solutions used for crystallization trials and LCP fluorescence recovery after photobleaching (FRAP) experiments.

Original languageEnglish (US)
Pages (from-to)342-349
Number of pages8
JournalMethods
Volume55
Issue number4
DOIs
StatePublished - Dec 1 2011
Externally publishedYes

Keywords

  • Lipidic cubic phase
  • Monoolein
  • Phase behavior
  • SAXS

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)

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