X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death

Muchmore, Steven W.; Sattler, Michael; Liang, Heng; Meadows, Robert P.; Harlan, John E.; Yoon, Ho Sup; Nettesheim, David G.; Chang, Brian M.; Thompson, Craig B.; Wong, Sui‐Lam; Ng, Shi-Chung; Fesik, Stephen W.

doi:10.1038/381335a0

Cited by 1,393 publications

(1,190 citation statements)

References 26 publications

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“…42 Early on, X-ray and NMR structural analysis revealed that an important inhibitor of PCD, BCL-x L , contains a 60-residue IDPR connecting helices al and a2. 43 Other members of the BCL-2 family, BH3-only proteins (such as BIM, BAD, and BMF) that serve as key initiators of PCD are largely disordered in solution. 44 Interaction of the disordered BIM, BAD, BMF, BAK, and tBID with several pro-survival BCL-2 family members leads to the formation of an a-helical segment that anchors these BH3-only proteins to the hydrophobic groove of their binding partners.…”

Section: Discussionmentioning

confidence: 99%

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

et al. 2013

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It is recognized now that intrinsically disordered proteins (IDPs), which do not have unique 3D structures as a whole or in noticeable parts, constitute a significant fraction of any given proteome. IDPs are characterized by an astonishing structural and functional diversity that defines their ability to be universal regulators of various cellular pathways. Programmed cell death (PCD) is one of the most intricate cellular processes where the cell uses specialized cellular machinery and intracellular programs to kill itself. This cell-suicide mechanism enables metazoans to control cell numbers and to eliminate cells that threaten the animal's survival. PCD includes several specific modules, such as apoptosis, autophagy, and programmed necrosis (necroptosis). These modules are not only tightly regulated but also intimately interconnected and are jointly controlled via a complex set of protein-protein interactions. To understand the role of the intrinsic disorder in controlling and regulating the PCD, several large sets of PCD-related proteins across 28 species were analyzed using a wide array of modern bioinformatics tools. This study indicates that the intrinsic disorder phenomenon has to be taken into consideration to generate a complete picture of the interconnected processes, pathways, and modules that determine the essence of the PCD. We demonstrate that proteins involved in regulation and execution of PCD possess substantial amount of intrinsic disorder. We annotate functional roles of disorder across and within apoptosis, autophagy, and necroptosis processes. Disordered regions are shown to be implemented in a number of crucial functions, such as protein-protein interactions, interactions with other partners including nucleic acids and other ligands, are enriched in post-translational modification sites, and are characterized by specific evolutionary patterns. We mapped the disorder into an integrated network of PCD pathways and into the interactomes of selected proteins that are involved in the p53-mediated apoptotic signaling pathway.

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Section: Discussionmentioning

confidence: 99%

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

et al. 2013

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“…These proteins share a similar helical bundle architecture composed of a hydrophobic, helical hairpin surrounded by a sheath of amphiphilic α-helices. Structurally, Bcl-x L ΔTM is most similar to the translocation domain of diphtheria toxin (23). For this toxin, as well as others, membrane association is mediated in part by the so-called hydrophobic, helical hairpin, which corresponds to α-helices 5 and 6 in Bcl-x L ΔTM (Figure 5a) (38)(39)(40)(41)(42)(43)(44)(45)(46).…”

Section: Two Acidic Residues In the Conserved Hairpin And The Conformmentioning

confidence: 99%

“…Because full-length Bcl-x L is anchored to the membrane by its transmembrane domain, experiments with this construct are not ideal for understanding the conformational change that primarily involves the N-terminal domain. Therefore, in this work, we describe experiments with synthetic lipid vesicles and Bcl-x L ΔTM, a construct that retains pro-survival activity in vivo (23). We determine the pH dependence of this process by testing for protein association with lipid vesicles as a function of pH with a sedimentation assay.…”

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The N-Terminal Domain of Bcl-x_L Reversibly Binds Membranes in a pH-Dependent Manner

et al. 2006

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Bcl-x L regulates apoptosis by maintaining the integrity of the mitochondrial outer membrane by adopting both soluble and membrane-associated forms. The membrane-associated conformation does not require a conserved, C-terminal transmembrane domain and appears to be inserted into the bilayer of synthetic membranes as assessed by membrane permeabilization and critical surface pressure measurements. Membrane association is reversible and is regulated by the cooperative binding of approximately two protons to the protein. Two acidic residues, Glu153 and Asp156, that lie in a conserved hairpin of Bcl-x L ΔTM appear to be important in this process on the basis of a 16% increase in the level of membrane association of the double mutant E153Q/D156N. Contrary to that for the wild type, membrane permeabilization for the mutant is not correlated with membrane association. Monolayer surface pressure measurements suggest that this effect is primarily due to less membrane penetration. These results suggest that E153 and D156 are important for the Bclx L ΔTM conformational change and that membrane binding can be distinct from membrane permeabilization. Taken together, these studies support a model in which Bcl-x L activity is controlled by reversible insertion of its N-terminal domain into the mitochondrial outer membrane. Future studies with Bcl-x L mutants such as E153Q/D156N should allow determination of the relative contributions of membrane binding, insertion, and permeabilization to the regulation of apoptosis.Bcl-2 proteins act as checkpoints in the regulation of apoptosis by integrating intracellular signals to control the permeability and possibly the morphology of the mitochon-drion (1-9). For many Bcl-2 proteins, this checkpoint involves a change in localization from the cytosol to the mitochondrion (10-13). For example, Bcl-x L displays mixed localization to the cytosol and to organellar membranes, including the mitochondrion. After an apoptotic stimulus, Bcl-x L appears to localize primarily to the mitochondrial outer membrane where it may bind other apoptotic factors such as Bad (10,11,14) or form an ion channel thought to maintain the integrity of the mitochondrial membrane (11,(15)(16)(17). While mixed localization of Bcl-x L is well-established, the relative importance of cytosolic-and membranous-localized protein to the regulation of apoptosis is not. Intriguingly, the membrane activity of Bcl-x L may contribute more to its anti-apoptotic activity than its ability to sequester pro-apoptotic proteins: a mutant

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“…Since Bcl-2 can in¯uence Ca ++ across membranes (Ba y et al, 1993;Lam et al, 1994), it has been also proposed that Bcl-2 or its family members may be involved in tra cking of ions (Reed 1994). In an analysis of the three-dimensional structure of Bcl-X L , Muchmore et al (1996) demonstrated signi®cant similarity with the pore-forming domains of bacterial toxins and proposed that Bcl-X L or its family members may be involved in regulating ion-channels across membranes. In support of this proposal, Schendel et al (1997) demonstrated the ability of recombinant Bcl-2 to form ion channels using a synthetic lipid bilayer-based assay.…”

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confidence: 99%

Nuclear targeting of Bax during apoptosis in human colorectal cancer cells

et al. 1998

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Homeostasis in colonic epithelial cells is regulated by the balance between proliferative activity and cell loss by apoptosis. Because epithelial cells at the apex of colonic crypts undergo apoptosis and proliferative activity is usually restricted to the base of the crypts, it has been proposed that the limited availability of growth factorsignals at the upper portions of the crypts may trigger apoptosis. In the present studies, we investigate the mechanism of apoptosis mediated by growth factor deprivation in colorectal carcinoma cells by delineating the possible involvement of Bax and its subcellular localization. We report that inhibition of epidermal growth factor receptor (EGFR) tyrosine kinase activity and downregulation of EGFR by anti-EGFR mAb 225 induces apoptosis in human colorectal carcinoma DiFi and FET cells. Induction of apoptosis was preceded by enhanced expression of newly synthesized Bax protein, and required protein synthesis. In the mAb 225-treated cells, Bax was redistributed from the cytosol to the nucleus and subsequently, to the nuclear membranes. The observed induction of Bax expression by mAb 225 was not associated with p53 induction. However, mAb 225 treatment also triggered relocalization of p53 from the cytosol to a nuclear membrane-bound form. Induction of Bax and its redistribution to the nucleus of DiFi cells during apoptosis was also demonstrated in response to butyrate, a physiological relevant molecule in colonic epithelial cells as it is the principal short-chain fatty acid produced by bacterial fermentation of dietary ®ber in colonic epithelium. Using immuno¯uorescence and confocal microscopy, we observed that Bax is predominantly localized in the cytosol, but during apoptosis it is localized both inside and along the nuclear membrane. Taken together, these ®ndings suggest that apoptosis induced by growth factor-deprivation or butyrate may involve the subcellular redistribution of Bax in human colorectal carcinoma cells.

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X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death

Cited by 1,393 publications

References 26 publications

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

The N-Terminal Domain of Bcl-x_L Reversibly Binds Membranes in a pH-Dependent Manner

Nuclear targeting of Bax during apoptosis in human colorectal cancer cells

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X-ray and NMR structure of human Bcl-xL, an inhibitor of programmed cell death

Cited by 1,393 publications

References 26 publications

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

Resilience of death: intrinsic disorder in proteins involved in the programmed cell death

The N-Terminal Domain of Bcl-xL Reversibly Binds Membranes in a pH-Dependent Manner

Nuclear targeting of Bax during apoptosis in human colorectal cancer cells

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The N-Terminal Domain of Bcl-x_L Reversibly Binds Membranes in a pH-Dependent Manner