Conformational States of the Cytoprotective Protein Bcl-xL

Abstract

The human protein Bcl-xL is a key regulator of programmed cell death in health and disease. Structural studies, important for understating the molecular basis for its functions, have advanced primarily by deleting both the long disordered loop that regulates its activity and the C-terminal tail that anchors the protein to intracellular membranes Here we describe the preparation and conformations of full-length Bcl-xL in both its water-soluble and membrane-anchored states. The study provides new biophysical insights about Bcl-xL and its greater Bcl-2 protein family.

Insight into Functional Membrane Proteins by Solution NMR: The Human Bcl-2 Protein—A Promising Cancer Drug Target

“…The Bcl-2 ∆TM construct with its hydrophobic TM-helix removed is partially soluble. This soluble escape (partial solubility) of Bcl-2 ∆TM can be explained based on the crystal structures of truncated soluble Bcl-2 variants where BH4 and BH3-1 domains are non-covalently packed while the FLD or truncated loop is extended to the outside of Bcl-2, and is also seen in other systems such as Bcl-xL [ 13 ]. Upon membrane insertion, the Bcl-2 domain organization based on the functions are shown as N-terminal BH4, FLD, and C-terminal BH3-1 and TM domains.…”

“…In addition, there are further challenges for structural determination for those proteins, since their functioning and interplay occur mainly in a membrane-embedded active state [ 8 , 10 , 11 ]. While structures exist for Bax in its soluble cytosolic state and soluble N-terminal nano-disc anchored Bcl-xL [ 12 , 13 ], there are no known atomic detail structures for the intact, insoluble human Bcl-2 protein in its membrane-embedded state, but only for a soluble, chimeric, and truncated version with 166 aa. For this Bcl-2 version an NMR-derived structure is available [ 14 ], and also crystal structures [ 15 ].…”

Effects of Cardiolipin on the Conformational Dynamics of Membrane-Anchored Bcl-xL

“…This requires the transition of Bcl-xL from an inactive state in the cytosol to an active state in the bilayer [10,11]. Two different membrane-integrated forms of Bcl-xL have been identified in vitro: (1) an anchored conformation in which the C-terminal helix of Bcl-xL serves as a transmembrane anchor, while the remaining helices retain their soluble fold (Figure 1A, red) [12][13][14], and (2) a membrane inserted conformation that involves the interaction of several Bcl-xL regions with the bilayer and extensive refolding, which leads to the release of the N-terminal BH4 helix (Figure 1A, blue) [15,16].…”

“…The specific roles of the different membrane-integrated conformations of Bcl-xL and their anti-apoptotic functions remain unclear; nevertheless, each has been linked to a different mode of apoptotic regulation. In the case of the anchored conformation, the lack of the refolding of the soluble head region (helices α1 through α7), as compared to the fold in the solution, results in an intact BH3-binding groove [12][13][14] (Figure 1B,C). BH3 domains are conserved regions in Bcl-xL homologs, such as its targets BAX and BAK, that are used to recognize and bind other apoptotic regulators [8,10,11].…”

“…Two different membrane-integrated Bcl-xL conformations have been identified: membrane-inserted and membrane-anchored Bcl-xL. The former is characterized by extensive refolding [15,16], while the latter retains the fold of the soluble state (aside from the released α8 anchor helix) [12][13][14]. Here, we focus on the membrane-anchored conformation of Bcl-xL and the influence of cardiolipin and protonation on its conformational dynamics.…”