Effect of phloretin on the binding of anilinin- naphtalen sulfonate to PC liposomes

CUTRO ANDREA; ROVERI OSCAR A.; MONTICH GUILLERMO

San Javier. Tucumásn

Congreso; XLI Reunión Anual de la Sociedad Argentina de Biofísica; 2012

sociedad argentina de biofísica

Phloretin, a known dipole potential modifier, uncoupled the mitochondrial ATP synthesis likely by modifying surface and/or structural properties of the inner mitochondrial membrane1. To test such hypothesis we studied its effect on the binding of anilin-naphtalen sulphonate (ANS) to liposomes. Phloretin almost completely inhibited (IC50=140 µM) the fluorescence increase observed when ANS binds to 1,2-dioleil-sn-glycerol-3 -phosphocholine (DOPC). ANS affinity was slightly decreased due to a lower binding entropy change. It also decreased the fluorescence quantum yield of the bound ANS and the number of binding sites. Phloretin modifies the thermotropic behavior of liposomes of 1,2-dimiristoil-phosphocholine (DMPC) decreasing its Tc from 24.8 to 17°C.Concerning the binding of ANS, results similar to those described in the previous paragraph were obtained with DMPC and 1,2-palmitoil-sn -glycerol-3 phosphocholine (DPPC) liposomes in liquid-crystalline phase. However, when the experiments were carried out at temperatures below Tc a clearly different behavior was observed. The fluorescence increase determined when ANS binds to DPPC liposomes was only slightly inhibited. The inhibition followed a sigmoidal response on [phloretin]. Conversely, the fluorescence increase when ANS binds to DMPC liposomes was 2.5 times higher in the presence of phloretin than in its absence (C0.5=71 µM). This last effect was mainly due to an increase in the affinity of ANS for the DMPC bilayer in the gel phase. No effect of phloretin on the fluorescence quantum yield was observed. In summary, phloretin binds to phospholipid bilayers in the liquidcrystalline and in the gel phases, exhibiting a higher affinity for the latter. On the basis of the results described here we postulate that it binds to DMPC bilayers in the gel phase increasing the surface potential not modifying the structural properties of the ANS binding sites, whereas it binds to the liquid-crystalline phase in a different manner modifying the water accessibility of the ANS binding sites. 1. Castelli, M, Fábregas, J, Roveri, O. XXXV Reunión de SAB Noviembre 2006, Rosario. Con subsidios de CONICET, UNR y ANPCyT.