Integralmembraneproteins(IMPs)insertintoBIOLOGicalmembranessuchthatthecellmembrane,whichisalipidbilayerapproximately30Åinthickness,shieldsthehydrophobicmembrane-spanningregionsoftheprotein.TopurifyIMPs,theproteinmustfirstbeextractedfromthebilayerthroughthereplacementofcelllipidswithsolubilizingdetergents.TheselectionofdetergentforboththeinitialsolubilizationoftheIMPandfordownstreamstudyisexceptionallyimportant,asonemustensurethattheselecteddetergentdoesnotinhibitfunction,causeirreversIBLedenaturationorinterferewithproteinpurificationorcrystallization.
PreviousstudiesontheselectionofdetergentsfortheinitialextractionofIMPshavedemonstratedtheimportanceofdetergentscreening(REFS).OneofthemostexhaustivestudieswasperformedbyWhiteetAl.(2007)inwhich122high-expressingpredictedmembraneproteinswerescreenedin6commonlyuseddetergentsTritonX-100,LDAO,FC-12,C8E4andDDM.Inthesetests,thezwitterionicdetergentsLDAOandFC-12werethemostefficientatsolubilizingyeastmembraneproteins;however,allofthedetergentsinthepanelsolubilizedatleast35%oftheproteinstested.
TheAnalyticExtractorkitcontainsapanelofeightdetergentscommonlyusedforthesolubilizationofIMPsfromthecellmembrane.Thedetergentsinthispanelwerechosenbasedontheirprioreffectivenessanduseintheliterature,andtheworkingconcentrationofeachdetergentisdependentonthecriticalmicelleconcentration(CMC)ofeachdetergent(i.e.higherconcentrationsoflowerCMCdetergentscomparedtohigherCMCdetergents)(ReftoWiener,2004).Theresultsfromthisassaywillaidintheselectionofadetergentforfurtherdownstreamstudies,includingx-raycrystallography,NMR,andbiochemicalassays.
