Two-dimensional electrophoresis of membrane proteins.
One third of all genes of various organisms encode membrane proteins, emphasizing their crucial cellular role. However, due to their high hydrophobicity, membrane proteins demonstrate low solubility and a high tendency for aggregation. Indeed, conventional two-dimensional gel electrophoresis (2-DE), a powerful electrophoretic method for the separation of complex protein samples that applies isoelectric focusing (IEF) in the first dimension and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) in the second dimension, has a strong bias against membrane proteins. This review describes two-dimensional electrophoretic techniques that can be used to separate membrane proteins. Alternative methods for performing conventional 2-DE are highlighted; these involve replacing the IEF with electrophoresis using cationic detergents, namely 16-benzyldimethyl-n-hexadecylammonium chloride (16-BAC) and cetyl trimethyl ammonium bromide (CTAB), or the anionic detergent SDS. Finally, the separation of native membrane protein complexes through the application of blue and clear native gel electrophoresis (BN/CN-PAGE) is reviewed, as well as the free-flow electrophoresis (FFE) of membranes.
Braun
Ralf J
RJ
GSF-National Research Center for Environment and Health, Institute of Human Genetics, Ingolstaedter Landstrasse 1, 85764, Munich-Neuherberg, Germany.
Kinkl
Norbert
N
Beer
Monika
M
Ueffing
Marius
M
D002458:Cell Fractionation / Q000379:methods; D003902:Detergents / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D007525:Isoelectric Focusing / Q000379:methods; D008565:Membrane Proteins / Q000032:analysis / Q000302:isolation & purification*; D020543:Proteome / Q000032:analysis / Q000302:isolation & purification; D040901:Proteomics / Q000379:methods*
D003902:Detergents; D008565:Membrane Proteins; D020543:Proteome