title: Unspecific membrane protein-lipid recognition: combination of AFM imaging, force spectroscopy, DSC and FRET measurements.
abstract: In this work, we will describe in quantitative terms the unspecific recognition between lactose permease (LacY) of Escherichia coli, a polytopic model membrane protein, and one of the main components of the inner membrane of this bacterium. Supported lipid bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) (3:1, mol/mol) in the presence of Ca(2+) display lateral phase segregation that can be distinguished by atomic force microscopy (AFM) as well as force spectroscopy. LacY shows preference for fluid (Lα) phases when it is reconstituted in POPE : POPG (3:1, mol/mol) proteoliposomes at a lipid-to-protein ratio of 40. When the lipid-to-protein ratio is decreased down to 0.5, two domains can be distinguished by AFM. While the upper domain is formed by self-segregated units of LacY, the lower domain is constituted only by phospholipids in gel (Lβ) phase. On the one hand, classical differential scanning calorimetry (DSC) measurements evidenced the segregation of a population of phospholipids and point to the existence of a boundary region at the lipid-protein interface. On the other hand, Förster Resonance Energy Transfer (FRET) measurements in solution evidenced that POPE is selectively recognized by LacY. A binary pseudophase diagram of POPE : POPG built from AFM observations enables to calculate the composition of the fluid phase where LacY is inserted. These results are consistent with a model where POPE constitutes the main component of the lipid-LacY interface segregated from the fluid bulk phase where POPG predominates.
authors.affiliation: Departament de Fisicoquímica, Facultat de Farmàcia and Institut de Nanociència i Nanotecnologia IN2UB, Barcelona, Catalonia, 08028, Spain.
authors.forename: Jordi H
authors.initials: JH
authors.lastname: Borrell
authors.affiliation: Departament de Fisicoquímica, Facultat de Farmàcia and Institut de Nanociència i Nanotecnologia IN2UB, Barcelona, Catalonia, 08028, Spain.
authors.forename: M Teresa
authors.initials: MT
authors.lastname: Montero
authors.affiliation: Unitat de Biofísica, Departament de Bioquímica i Biología Molecular, Facultat de Medicina UAB, Bellaterra (Barcelona), 08193, Spain.
authors.forename: Antoni
authors.initials: A
authors.lastname: Morros
authors.affiliation: Departament de Fisicoquímica, Facultat de Farmàcia and Institut de Nanociència i Nanotecnologia IN2UB, Barcelona, Catalonia, 08028, Spain.
authors.forename: Òscar
authors.initials: Ò
authors.lastname: Domènech
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D002152:Calorimetry, Differential Scanning / Q000379:methods; D004926:Escherichia coli / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D008051:Lipid Bilayers / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism; D018625:Microscopy, Atomic Force / Q000379:methods; D010715:Phosphatidylglycerols / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D011510:Proteolipids / Q000378:metabolism; D013057:Spectrum Analysis / Q000379:methods
chemical_list: D001426:Bacterial Proteins; D008051:Lipid Bilayers; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D010715:Phosphatidylglycerols; D010743:Phospholipids; D011510:Proteolipids; C020485:proteoliposomes; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol; C021499:lactose permease
keywords: AFM; DSC; FRET; FS; lactose permease; lipid-protein interaction; phsopholipids