tBid Undergoes Multiple Conformational Changes at the Membrane Required for Bax Activation |
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Authors: | Aisha Shamas-Din Scott Bindner Weijia Zhu Yehudit Zaltsman Clinton Campbell Atan Gross Brian Leber David W. Andrews Cécile Fradin |
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Affiliation: | From the ‡Department of Biochemistry and Biomedical Sciences and ;¶Department of Medicine, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.;the §Department of Biological Regulation, The Weizmann Institute of Science, Rehovot 76100, Israel, and ;the ‖Department of Physics and Astronomy, McMaster University, Hamilton, Ontario L8S 4M1, Canada |
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Abstract: | Bid is a Bcl-2 family protein that promotes apoptosis by activating Bax and eliciting mitochondrial outer membrane permeabilization (MOMP). Full-length Bid is cleaved in response to apoptotic stimuli into two fragments, p7 and tBid (p15), that are held together by strong hydrophobic interactions until the complex binds to membranes. The detailed mechanism(s) of fragment separation including tBid binding to membranes and release of the p7 fragment to the cytoplasm remain unclear. Using liposomes or isolated mitochondria with fluorescently labeled proteins at physiological concentrations as in vitro models, we report that the two components of the complex quickly separate upon interaction with a membrane. Once tBid binds to the membrane, it undergoes slow structural rearrangements that result in an equilibrium between two major tBid conformations on the membrane. The conformational change of tBid is a prerequisite for interaction with Bax and is, therefore, a novel step that can be modulated to promote or inhibit MOMP. Using automated high-throughput image analysis in cells, we show that down-regulation of Mtch2 causes a significant delay between tBid and Bax relocalization in cells. We propose that by promoting insertion of tBid via a conformational change at the mitochondrial outer membrane, Mtch2 accelerates tBid-mediated Bax activation and MOMP. Thus the interaction of Mtch2 and tBid is a potential target for therapeutic control of Bid initiated cell death. |
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Keywords: | Apoptosis Bax Bcl-2 Proteins Fluorescence Correlation Spectroscopy Fluorescence Resonance Energy Transfer (FRET) Protein Conformation Bid Mtch2 Fluorescence Spectroscopy tBid |
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