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Preparation of microcrystals in lipidic cubic phase for serial femtosecond crystallography

Academic Article
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Overview

authors

  • Liu, W.
  • Ishchenko, A.
  • Cherezov, Vadim

publication date

  • September 2014

journal

  • Nature Protocols  Journal

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.

subject areas

  • Crystallography
  • Lasers
  • Lipid Bilayers
  • Membrane Proteins
  • Time Factors
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Identity

PubMed Central ID

  • PMC4209290

International Standard Serial Number (ISSN)

  • 1754-2189

Digital Object Identifier (DOI)

  • 10.1038/nprot.2014.141

PubMed ID

  • 25122522
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Additional Document Info

start page

  • 2123

end page

  • 2134

volume

  • 9

issue

  • 9

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