Membrane protein separation and analysis by supercritical fluid chromatography-mass spectrometry.
Membrane proteins comprise 25-30% of the human genome and play critical roles in a wide variety of important biological processes. However, their hydrophobic nature has compromised efforts at structural characterization by both X-ray crystallography and mass spectrometry. The detergents that are generally used to solubilize membrane proteins interfere with the crystallization process essential to X-ray studies and cause severe ion suppression effects that hinder mass spectrometric analysis. In this report, the use of supercritical fluid chromatography-mass spectrometry for the separation and analysis of integral membrane proteins and hydrophobic peptides is investigated. It is shown that detergents are rapidly and effectively separated from the proteins and peptides, yielding them in a state suitable for direct mass spectrometric analysis.
Zhang
Xu
X
Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, Wisconsin 53706, USA.
Scalf
Mark
M
Westphall
Michael S
MS
Smith
Lloyd M
LM
D001436:Bacteriorhodopsins / Q000032:analysis* / Q000378:metabolism; D025924:Chromatography, Supercritical Fluid / Q000295:instrumentation* / Q000379:methods*; D013058:Mass Spectrometry / Q000295:instrumentation* / Q000379:methods*; D045332:Photosystem II Protein Complex / Q000032:analysis / Q000378:metabolism; D011485:Protein Binding; D018724:Spinacia oleracea / Q000201:enzymology
D045332:Photosystem II Protein Complex; D001436:Bacteriorhodopsins