title: Phosphatidylethanolamine-lactose permease interaction: a comparative study based on FRET.
abstract: In this work we have investigated the selectivity of lactose permease (LacY) of Escherichia coli (E. coli) for its surrounding phospholipids when reconstituted in binary mixtures of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE), 1,2-Palmitoyl-sn-glycero-3-phosphoethanolamine (DPPE), or 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) with 1-palmitoyl-2-oleoyl-sn-glycero-3-(phospho-rac-(1-glycerol)) (POPG). Förster resonance energy transfer (FRET) measurements have been performed to investigate the selectivity between a single tryptophan mutant of LacY used as donor (D), and two analogues of POPE and POPG labeled with pyrene in the acyl chains (Pyr-PE and Pyr-PG) used as acceptors. As a difference from previous works, now the donor has been single-W151/C154G/D68C LacY. It has been reported that the replacement of the aspartic acid in position 68 by cysteine inhibits active transport in LacY. The objectives of this work were to elucidate the phospholipid composition of the annular region of this mutant and to determine whether the mutation performed, D68C, induced changes in the protein-lipid selectivity. FRET efficiencies for Pyr-PE were always higher than for Pyr-PG. The values of the probability of each site in the annular ring being occupied by a label (μ) were similar at the studied temperatures (24 °C and 37 °C), suggesting that the lipid environment is not significantly affected when increasing the temperature. By comparing the results with those obtained for single-W151/C154G LacY, we observe that the mutation in the 68 residue indeed changes the selectivity of the protein for the phospholipids. This might be probably due to a change in the conformational dynamics of LacY.
authors.affiliation: Departament de Fisicoquímica, Facultat de Farmàcia, Universitat de Barcelona, Spain.
authors.forename: Carme
authors.initials: C
authors.lastname: Suárez-Germà
authors.forename: Luís M S
authors.initials: LM
authors.lastname: Loura
authors.forename: Oscar
authors.initials: O
authors.lastname: Domènech
authors.forename: M Teresa
authors.initials: MT
authors.lastname: Montero
authors.forename: José Luís
authors.initials: JL
authors.lastname: Vázquez-Ibar
authors.forename: Jordi
authors.initials: J
authors.lastname: Hernández-Borrell
mesh_terms: D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D008958:Models, Molecular; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000378:metabolism*; D010715:Phosphatidylglycerols / Q000378:metabolism; D017354:Point Mutation; D027981:Symporters / Q000235:genetics / Q000378:metabolism*
chemical_list: C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D009004:Monosaccharide Transport Proteins; D010714:Phosphatidylethanolamines; D010715:Phosphatidylglycerols; D027981:Symporters; C057561:1-palmitoyl-2-oleoylphosphatidylethanolamine; C007510:1,2-dipalmitoyl-3-phosphatidylethanolamine; C483858:phosphatidylethanolamine; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol