title: Interaction of Bestrophin-1 with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) in surface films.
abstract: Human bestrophin-1 (hBest1) is a transmembrane channel protein, predominantly expressed in the membrane of retinal pigment epithelium (RPE) cells. Although it is clear that hBest1's interactions with lipids are crucial for its function such studies were not performed as the protein was not purified. Here we describe an effective purification of hBest1 from Madin-Darby Canine Kidney (MDCK) cells via simple gel-filtration and affinity chromatographic steps, which makes possible to probe the protein interplay with lipids. The interaction of the purified hBest1 with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) was studied in Langmuir monolayers. The surface pressure (π)-area (A) isotherms and compression/expansion isocycles of POPC monolayer were recorded in absence and presence of hBest1 in the subphase. The π(A) isotherms were analyzed in terms of surface compressional modulus and via two-dimensional virial equation of state. The dilatational rheological properties of the surface films and their surface potential were also measured. The morphology of the films was observed by Brewster angle microscopy. The inclusion of the protein in the film subphase does not lead to in-depth penetration of hBest1 but interaction takes place in the headgroup region of the monolayer. The hBest1/POPC interaction resulted in formation of more condensed films, which rheological properties and lateral structure differed significantly from the pure POPC monolayers. Our study sheds light on the still unclear question how hBest1 gets in touch with biomembrane phospholipids of eukaryotic cells that might be of key importance for the proper structure and function of RPE biomembranes.
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Biochemistry, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria.
authors.forename: Kirilka
authors.initials: K
authors.lastname: Mladenova
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Biochemistry, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria.
authors.forename: Svetla D
authors.initials: SD
authors.lastname: Petrova
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Biochemistry, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria.
authors.forename: Georgi As
authors.initials: GA
authors.lastname: Georgiev
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Cytology, Histology and Embryology, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria.
authors.forename: Veselina
authors.initials: V
authors.lastname: Moskova-Doumanova
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Biochemistry, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria.
authors.forename: Zdravko
authors.initials: Z
authors.lastname: Lalchev
authors.affiliation: Sofia University "St. Kliment Ohridski", Department of Biochemistry, Faculty of Biology, 8 Dragan Tzankov str., 1164 Sofia, Bulgaria. Electronic address: doumanov@biofac.uni-sofia.bg.
authors.forename: Jordan A
authors.initials: JA
authors.lastname: Doumanov
mesh_terms: D000818:Animals; D000075002:Bestrophins; D015153:Blotting, Western; D018118:Chloride Channels / Q000302:isolation & purification / Q000378:metabolism*; D002850:Chromatography, Gel; D004285:Dogs; D005136:Eye Proteins / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D061985:Madin Darby Canine Kidney Cells; D010713:Phosphatidylcholines / Q000378:metabolism*; D011485:Protein Binding; D013499:Surface Properties
chemical_list: C113133:BEST1 protein, human; D000075002:Bestrophins; D018118:Chloride Channels; D005136:Eye Proteins; D010713:Phosphatidylcholines; C028694:1-palmitoyl-2-oleoylphosphatidylcholine
keywords: BAM; Langmuir monolayers; Lipid–protein interactions; MDCK cells; POPC; hBest1