Cardiolipin, phosphatidylserine, and BH4 domain of Bcl-2 family regulate Ca2+/H+ antiporter activity of human Bax inhibitor-1.

We investigated the effects of phospholipid composition in membranes and Bcl-2 homology (BH) domains of the Bcl-2 family on Ca2+/H+ antiporter activity of human recombinant Bax inhibitor-1 (BI-1) reconstituted into membranes. Cardiolipin (CL) and phosphatidylserine (PS) stimulated the proton-mediated efflux of Ca2+ ions encapsulated into proteoliposomes when compared to Ca2+ efflux from 100% phosphatidylcholine (PC) membranes in a CL or PS concentration-dependent manner. Concomitantly, the anionic phospholipids also enhanced H+ ion influx into the membranes. Lateral segregations of CL and PS were observed through the fluorescence properties of fluorophore-labeled phospholipids upon BI-1 reconstitution in PC/CL or PC/PS binary systems. However, other anionic phospholipids, such as phosphatidic acid, phosphatidylglycerol, and phosphatidylinositol did not influence the stimulation of BI-1 functions in membranes. The peptide corresponding to the BH4 domain of Bcl-2 and Bcl-xL proteins stimulated the BI-1 activities in 100% PC membranes. The peptide also showed an additive effect with CL or PS. Furthermore, the CL, PS, and BH4 domains specifically increased oligomerization levels such as dimer and tetramer of BI-1 in membranes. Taken together, these results suggest that the CL, PS, and BH4 domains were stimulating factors for the Ca2+/H+ antiporter activities of BI-1 through protein oligomerization.