The Role of N-Terminal Domain Exposure of Bax in the Regulation of Intrinsic Apoptosis at the Mitochondria
The Bcl-2 protein family, made up of pro and anti-apoptotic proteins plays a major role in regulating apoptosis. Bax is a pro-apoptotic member of the Bcl-2 family. In its inactive form, Bax is primarily a cytosolic and monomeric protein. Upon activation, Bax translocates to and permeabilizes the mitochondrial outer membrane releasing apoptotic factors. Because Bax plays such a pivotal role in cell death signaling, it is highly regulated through interaction with protein kinases and other members of the Bcl-2 family. Activation of Bax also requires a substantial change in conformation and of localization of the Bax protein, exposure of the N-terminus is one such conformational change Bax undergoes. However, the molecular processes linking N-terminus exposure with respect to Bax’s ability to translocate to, insert, and permeabilize the mitochondrial membrane remain to be defined. When Bax mutants P168A and S184A are expressed in yeast, they have higher mitochondrial localization than WT Bax. However, The S184A mutant is far less capable than P168A or WT Bax in releasing cytochrome c, a step considered to be the point of no return in the apoptotic signaling cascade. By using an ELISA to probe the recombinant Bax mutants for N-terminus exposure, we found significantly lower levels of N-terminus exposure the S184A when compared to P168A or WT Bax, suggesting that the N-terminus plays a more predominant role in pore formation than mitochondrial localization. However, the mechanism by which the N terminus increases the pore formation ability of Bax has yet to be defined.