Band 3(B3),theaniontransporter,isanintegralmembraneproteinthatplaysakeystructuralroleby anchoringtheplasmamembranetothespectrin-basedmembraneskeletonintheredcell.Inaddition, it alsoplaysacriticalroleintheassemblyofglycolyticenzymestoregulateredcellmetabolism. However,itsabilitytorecruitproteinsthatcanpreventmembraneoxidationhasnotbeenpreviously explored.Inthisstudy,usingavarietyofexperimentalapproachesincludingcross-linkingstudies, fluorescenceanddichroicmeasurements,surfaceplasmonresonanceanalysis,andproteolyticdigestion assays,wedocumentthattheantioxidantproteinperoxiredoxin-2(PRDX2),thethirdmostabundant cytoplasmicproteininRBCs,interactswiththecytoplasmicdomainofB3.Thesurfaceelectrostatic potentialanalysisandstoichiometrymeasurementsrevealedthattheN-terminalpeptideofB3is involvedintheinteraction.PRDX2underwentaconformationalchangeuponitsbindingtoB3without losingitsperoxidaseactivity.HemichromeformationinducedbyphenylhydrazineofRBCsprevented membraneassociationofPRDX2,implyingoverlappingbindingsites.Documentationoftheabsenceof bindingofPRDX2toB3Neapolisredcellmembranes,inwhichtheinitialN-terminal11aminoacidsare deleted,enabledustoconcludethatPRDX2bindstotheN-terminalcytoplasmicdomainofB3andthat the first11aminoacidsofthisdomainarecrucialforPRDX2membraneassociationinintactRBCs. ThesefindingsimplyyetanotherimportantroleforB3inregulatingredcellmembranefunction.
Membrane association of peroxiredoxin-2 in red cells is mediated by the N-terminal cytoplasmic domain of band3
MATTE', Alessandro;BERTOLDI, Mariarita;SICILIANO, Angela;DE FRANCESCHI, Lucia
2013-01-01
Abstract
Band 3(B3),theaniontransporter,isanintegralmembraneproteinthatplaysakeystructuralroleby anchoringtheplasmamembranetothespectrin-basedmembraneskeletonintheredcell.Inaddition, it alsoplaysacriticalroleintheassemblyofglycolyticenzymestoregulateredcellmetabolism. However,itsabilitytorecruitproteinsthatcanpreventmembraneoxidationhasnotbeenpreviously explored.Inthisstudy,usingavarietyofexperimentalapproachesincludingcross-linkingstudies, fluorescenceanddichroicmeasurements,surfaceplasmonresonanceanalysis,andproteolyticdigestion assays,wedocumentthattheantioxidantproteinperoxiredoxin-2(PRDX2),thethirdmostabundant cytoplasmicproteininRBCs,interactswiththecytoplasmicdomainofB3.Thesurfaceelectrostatic potentialanalysisandstoichiometrymeasurementsrevealedthattheN-terminalpeptideofB3is involvedintheinteraction.PRDX2underwentaconformationalchangeuponitsbindingtoB3without losingitsperoxidaseactivity.HemichromeformationinducedbyphenylhydrazineofRBCsprevented membraneassociationofPRDX2,implyingoverlappingbindingsites.Documentationoftheabsenceof bindingofPRDX2toB3Neapolisredcellmembranes,inwhichtheinitialN-terminal11aminoacidsare deleted,enabledustoconcludethatPRDX2bindstotheN-terminalcytoplasmicdomainofB3andthat the first11aminoacidsofthisdomainarecrucialforPRDX2membraneassociationinintactRBCs. ThesefindingsimplyyetanotherimportantroleforB3inregulatingredcellmembranefunction.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.