Cancer arises as a result of the inability of damaged cells to initiate a signaling cascade that results in the destruction of the cell and clearance from the body. This process, known as apoptosis, is necessary to prevent reproduction of damaged cells that leads to cancer. A family of proteins known as the Bcl-2 protein family regulates apoptosis. The Bcl-2 protein family works to promote cell death through the interaction of death promoting proteins. Bax, a pro-apoptotic member of this family, initiates the cell death cascade by releasing cytochrome C. Bax has the ability to mediate cytochrome c release by working in conjunction with cardiolipin. Previous studies have shown that the presence of cardiolipin enhances the ability of Bax to form pores in reconstituted membranes through an, as yet, unknown mechanism. Preliminary data from our lab has begun to elucidate the mechanism of interaction between Bax and cardiolipin. In the presence of cardiolipin Bax undergoes a unique conformational change that facilitates the release of molecules from model membranes. The goals of the current project are to determine the impact that this conformational change has on the active and functional conformation of Bax. Our goals are to detail the structural changes and to correlate these changes with the function of Bax at the membrane. This will be achieved by studying pore formation and oligomerization of membrane-associated Bax using fluorescent spectroscopic techniques. We will also study how regions of Bax contribute to membrane recognition pore formation and oligomerization through the use of mutants. The two regions of Bax that are of interest are the BH3 domain, also called the putative cardiolipin binding region, and the c-terminal hydrophobic domain. Understanding the mechanisms of pore formation and oligomerization as well as how domains of Bax contribute to the membrane-active conformation will enhance our understanding of apoptosis and aid in our ability to overcome the barriers that cancer cells erect to evade apoptosis.
