Microscale NMR Screening of New Detergents for Membrane Protein Structural Biology

Qinghai Zhang†, Reto Horst†, Michael Geralt†, Xingquan Ma†, Wen-Xu Hong†, M. G. Finn‡§, Raymond C. Stevens†‡ and Kurt Wthrich†‡§
Department of Molecular Biology, Department of Chemistry, Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037
J. Am. Chem. Soc., 2008, 130 (23), pp 7357–7363
DOI: 10.1021/ja077863d
Publication Date (Web): May 14, 2008
Copyright © 2008 American Chemical Society
, †

Department of Molecular Biology

, ‡

Department of Chemistry.

, §

Skaggs Institute for Chemical Biology.

Abstract

Abstract Image

The rate limiting step in biophysical characterization of membrane proteins is often the availability of suitable amounts of protein material. It was therefore of interest to demonstrate that microcoil nuclear magnetic resonance (NMR) technology can be used to screen microscale quantities of membrane proteins for proper folding in samples destined for structural studies. Micoscale NMR was then used to screen a series of newly designed zwitterionic phosphocholine detergents for their ability to reconstitute membrane proteins, using the previously well characterized β-barrel E. coli outer membrane protein OmpX as a test case. Fold screening was thus achieved with microgram amounts of uniformly 2H,15N-labeld OmpX and affordable amounts of the detergents, and prescreening with SDS-gel electrophoresis ensured efficient selection of the targets for NMR studies. A systematic approach to optimize the phosphocholine motif for membrane protein refolding led to the identification of two new detergents, 138-Fos and 179-Fos, that yield 2D [15N,1H]-TROSY correlation NMR spectra of natively folded reconstituted OmpX.

View: Full Text HTML | Hi-Res PDF | PDF w/ Links

Tools

SciFinder Links

SciFinder subscribers: Click to sign in | Not a SciFinder subscriber? Learn more at www.cas.org

Explore by:

History

  • Published In Issue June 11, 2008
  • Article ASAPMay 14, 2008
  • Received: October 16, 2007

Recommend & Share

Related Content

Other ACS content by these authors: