A gas chromatography-mass spectrometry method to monitor detergents removal from a membrane protein sample.
In membrane protein biochemical and structural studies, detergents are used to mimic membrane environment and maintain functional, stable conformation of membrane proteins in the absence of lipid bilayers. However, detergent concentration, esp. molar ratio of membrane protein to detergent is usually unknown. Here, a gas chromatography-mass spectrometry selected ion monitoring (GC-MS-SIM) method was developed to quantify four detergents which are frequently used in membrane protein structural studies. To remove excessive detergents, a filtered centrifugation using Centricon tubes was applied. A membrane protein Ig-Beta fragment in four different detergent micelles was exemplified. Detergent concentrations in the upper and lower fraction of the Centricon tube were measured after each round of centrifugation. The results were very consistent to basic properties of detergent micelles in aqueous solvents. Therefore, coupling of GC-MS-SIM and detergent removal by Centricon tubes, detergents concentration, esp. molar ratio of membrane protein to detergent could be controlled, which will expedite membrane protein structural and biochemical studies.
Shi
Chaowei
C
National Laboratory for Physical Science at Microscale, School of Life Science, University of Science and Technology of China, 96 Jin Zhai Road, Hefei, Anhui 230026, People's Republic of China.
Han
Fang
F
Xiong
Ying
Y
Tian
Changlin
C
D003902:Detergents / Q000302:isolation & purification*; D004123:Dimethylamines / Q000302:isolation & purification; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D007136:Immunoglobulins / Q000032:analysis / Q000737:chemistry; D008565:Membrane Proteins / Q000032:analysis / Q000737:chemistry*; D012044:Regression Analysis
D003902:Detergents; D004123:Dimethylamines; D007136:Immunoglobulins; D008565:Membrane Proteins; C014518:dodecyldimethylamine oxide