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Miscibility of hBest1 and sphingomyelin in surface films – a prerequisite for interaction with membrane domains

  • Human bestrophin-1 (hBest1) is a transmembrane Ca2+- dependent anion channel, associated with the transport of Cl−, HCO3- ions, γ-aminobutiric acid (GABA), glutamate (Glu), and regulation of retinal homeostasis. Its mutant forms cause retinal degenerative diseases, defined as Bestrophinopathies. Using both physicochemical - surface pressure/mean molecular area (π/A) isotherms, hysteresis, compressibility moduli of hBest1/sphingomyelin (SM) monolayers, Brewster angle microscopy (BAM) studies, and biological approaches - detergent membrane fractionation, Laurdan (6-dodecanoyl-N,N-dimethyl-2-naphthylamine) and immunofluorescence staining of stably transfected MDCK-hBest1 and MDCK II cells, we report: 1) Ca2+, Glu and GABA interact with binary hBest1/SM monolayers at 35 °C, resulting in changes in hBest1 surface conformation, structure, self-organization and surface dynamics. The process of mixing in hBest1/SM monolayers is spontaneous and the effect of protein on binary films was defined as “fluidizing”, hindering the phase-transition of monolayer from liquid-expanded to intermediate (LE-M) state; 2) in stably transfected MDCK-hBest1 cells, bestrophin-1 was distributed between detergent resistant (DRM) and detergent-soluble membranes (DSM) - up to 30 % and 70 %, respectively; in alive cells, hBest1 was visualized in both liquid-ordered (Lo) and liquid-disordered (Ld) fractions, quantifying protein association up to 35 % and 65 % with Lo and Ld. Our results indicate that the spontaneous miscibility of hBest1 and SM is a prerequisite to diverse protein interactions with membrane domains, different structural conformations and biological functions.

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Author of HS ReutlingenAndreeva, Tonya
Erschienen in:Colloids and surfaces : an international journal devoted to fundamental and applied research on colloid and interfacial phenomena in relation to systems of biological origin. B, Biointerfaces
Place of publication:Amsterdam
Document Type:Journal article
Publication year:2020
Tag:MDCK II cells; hBest1; laurdan; lipid rafts; sphingomyelin; surface films
Page Number:9
First Page:1
Last Page:9
Article Number:110893
DDC classes:540 Chemie
Open access?:Nein
Licence (German):License Logo  In Copyright - Urheberrechtlich geschützt