Conformation of the Membrane-Integrated Functional State of Anti-Apoptotic BCL-XL

Abstract

BCL-XL is a major suppressor of apoptosis and a prominent member of the BCL-2 family. It is expressed in a variety of tissues and cell types and overexpressed in many tumors where it acts to promote tumor cell survival, tumor formation and tumor resistance to chemotherapy. Structure and activity studies have focused on truncated forms of the protein without the hydrophobic C-terminus that is essential for mediating the protein's association with intracellular membranes and cytoprotective activity. Hence, while the cytosolic state of the protein is relatively well understood, molecular information about the membrane-bound state is limited. We have used solution and solid-state NMR, together with isothermal titration calorimetry, to examine the structure and ligand binding activity of membrane-integrated BCL-XL, including its complete hydrophobic C-terminus. The fraction of BCL-XL expressed in E. coli as insoluble inclusion bodies can be isolated, purified and refolded into phospholipid bilayers. Here we describe the conformation of this membrane-bound state as a soluble globular domain anchored by a transmembrane C-terminal [[Unsupported Character - Symbol Font α]] helix. Membrane-bound BCL-XL binds a BID BH3 peptide with affinity similar to or slightly greater than the C-terminal truncated form. Implications for apoptosis from the conformation the membrane-integrated BCL-XL are discussed.Acknowledgements. This research was supported by grants from the National Institutes of Health (CA 179087; EB 002031).