Activity of phosphatidylinositol transfer protein is sensitive to ethanol and membrane curvature

Phosphatidylinositol transfer protein (PITP) is critical for many cellular signalling and trafficking events that are influenced by ethanol. The influ ence of ethanol and membrane curvature on the activity of recombinant mouse PITP-alpha in vitro is evaluated by monitoring the transfer of phosphatidy linositol (PtdIns) from rat hepatic microsomes to unilamellar vesicles. Acu te exposure to pharmacological levels of ethanol enhanced the function of P ITP. Chloroform shared a similar ability to enhance function when both drug concentrations were normalized to their respective octanol/water partition coefficients, indicating that the effect is not unique to ethanol and migh t be common to hydrophobic solutes. Neither the PITP activity nor its ethan ol enhancement was altered by using thermally pretreated (denatured) or pro tease-treated microsomes, indicating that the native microsomal protein str ucture was unlikely to be a determinant of transfer. Kinetic analyses indic ated that ethanol acted by increasing the PITP-mediated flux of PtdIns from both microsomal and liposomal surfaces. The activity of PITP was strongly dependent on the lipid structure, with a steep dependence on the expressed curvature of the membrane. Activity was greatest for small, highly curved s onicated vesicles and decreased markedly for large, locally planar unilamel lar vesicles. Ethanol enhanced PITP-mediated PtdIns transfer to all vesicle s, but its effect was much smaller than the enhancement due to curvature, w hich is consistent with ethanol's comparatively modest ability to perturb m embrane lipids. The ethanol efficacy observed is as pronounced as any previ ously described lipid-mediated ethanol action. In addition, these observati ons raise the possibility that PITP specifically delivers PtdIns to metabol ically active membrane domains of convex curvature and/or low surface densi ties of lipid.