Effective removal of nonionic detergents in protein mass spectrometry, hydrogen/deuterium exchange, and proteomics.
Detergents are frequently used for protein isolation and solubilization. Their presence is crucial in membrane protein protocols or in lipid raft proteomics. However, they are usually poorly compatible with mass spectrometry. Several different sample preparation protocols are routinely used, but they are either laborious or suffer from sample losses. Here, we describe our alternative method for nonionic detergent removal. It is based on selective detergent extraction after capture of the sample on a reversed phase cartridge. The extraction is performed by chlorinated solvents and works well for polyoxyethylene based nonionic detergents, but also for polymers like polyethylene and propylene glycol. Detergent removal can be also carried out on the protein level but a special care must be taken with hydrophobic proteins. In such cases, it is preferable to perform detergent removal after proteolysis which digests the protein to peptides and reduces the hydrophobicity. The method can easily be automated and is compatible with hydrogen/deuterium exchange coupled to mass spectrometry.
Rey
Martial
M
Laboratoire de Biochimie et Biophysique des Systèmes Intégrés (BBSI), Institut de Recherches en Technologies et Sciences pour le Vivant (iRTSV), UMR 5092 CNRS-CEA-UJF, Grenoble, F-38054, France.
Mrázek
Hynek
H
Pompach
Petr
P
Novák
Petr
P
Pelosi
Ludovic
L
Brandolin
Gérard
G
Forest
Eric
E
Havlícek
Vladimír
V
Man
Petr
P
D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods; D003902:Detergents / Q000302:isolation & purification*; D057927:Hydrophobic and Hydrophilic Interactions; D013058:Mass Spectrometry / Q000379:methods*; D008565:Membrane Proteins / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D040901:Proteomics / Q000379:methods; D012710:Serum Albumin, Bovine / Q000737:chemistry
D003902:Detergents; D008565:Membrane Proteins; D011506:Proteins; D012710:Serum Albumin, Bovine