Small-angle X-ray and neutron scattering demonstrates that cell-free expression produces properly formed disc-shaped nanolipoprotein particles

Thomas E. Cleveland, Wei He, Angela C. Evans, Nicholas O Fischer, Edmond Y Lau, Matthew A Coleman, Paul Butler

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

Nanolipoprotein particles (NLPs), composed of membrane scaffold proteins and lipids, have been used to support membrane proteins in a native-like bilayer environment for biochemical and structural studies. Traditionally, these NLPs have been prepared by the controlled removal of detergent from a detergent-solubilized protein-lipid mixture. Recently, an alternative method has been developed using direct cell-free expression of the membrane scaffold protein in the presence of preformed lipid vesicles, which spontaneously produces NLPs without the need for detergent at any stage. Using SANS/SAXS, we show here that NLPs produced by this cell-free expression method are structurally indistinguishable from those produced using detergent removal methodologies. This further supports the utility of single step cell-free methods for the production of lipid binding proteins. In addition, detailed structural information describing these NLPs can be obtained by fitting a capped core-shell cylinder type model to all SANS/SAXS data simultaneously.

Original languageEnglish (US)
Pages (from-to)780-789
Number of pages10
JournalProtein Science
Volume27
Issue number3
DOIs
StatePublished - Mar 1 2018
Externally publishedYes

Keywords

  • cell-free expression
  • nanodiscs
  • nanolipoprotein particles
  • NLPs
  • SANS
  • SAXS
  • small-angle scattering

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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