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How does the Bax-α1 targeting sequence interact with mitochondrial membrane?: The role of cardiolipin
Umeå University, Faculty of Science and Technology, Department of Chemistry. (UMR 5248 CBMN, CNRS, Université Bordeaux 1, ENITAB, IECB, 33607 Pessac Cedex, France)
UMR 5248 CBMN, CNRS, Université Bordeaux 1, France.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
2009 (English)In: Biochimica et Biophysica Acta - Biomembranes, ISSN 0005-2736, E-ISSN 1879-2642, Vol. 1788, no 3, 623-631 p.Article in journal (Refereed) Published
Abstract [en]

A key event in programmed cell death is the translocation of the apoptotic Bax protein from the cytosol towards mitochondria. The first helix localized at the N-terminus of Bax (Bax-α1) can act here as an addressing sequence, which directs activated Bax towards the mitochondrial surface. Solid state NMR (nuclear magnetic resonance), CD (circular dichroism) and ATR (attenuated total reflection) spectroscopy were used to elucidate this recognition process of a mitochondrial membrane system by Bax-α1. Two potential target membranes were studied, with the outer mitochondrial membrane (OM) mimicked by neutral phospholipids, while mitochondrial contact sites (CS) contained additional anionic cardiolipin. 1H and 31P magic angle spinning (MAS) NMR revealed Bax-α1 induced pronounced perturbations in the lipid headgroup region only in presence of cardiolipin. Bax-α1 could not insert into CS membranes but at elevated concentrations it inserted into the hydrophobic core of cardiolipin-free OM vesicles, thereby adopting β-sheet-like features, as confirmed by ATR. CD studies revealed, that the cardiolipin mediated electrostatic locking of Bax-α1 at the CS membrane surface promotes conformational change into an α-helical state; a process which seems to be necessary to induce further conformational transition events in activated Bax which finally causes irreversible membrane permeabilization during the mitochondrial apoptosis.

Place, publisher, year, edition, pages
Elsevier B.V. , 2009. Vol. 1788, no 3, 623-631 p.
Keyword [en]
Apoptosis, Bax N-terminal, Cardiolipin, Solid-State NMR
URN: urn:nbn:se:umu:diva-11590DOI: 10.1016/j.bbamem.2008.12.014OAI: diva2:151261
Available from: 2009-01-20 Created: 2009-01-20 Last updated: 2012-08-15Bibliographically approved
In thesis
1. Apoptosis Regulation via the Mitochondrial Pathway: Membrane Response upon Apoptotic Stimuli
Open this publication in new window or tab >>Apoptosis Regulation via the Mitochondrial Pathway: Membrane Response upon Apoptotic Stimuli
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The aim of this thesis was the investigation of the mitochondrial response mechanisms upon apoptotic stimuli. The specific objectives were the biophysical characterization of membrane dynamics and the specific roles of lipids in the context of apoptotic regulation occurring at the mitochondrion and its complex membrane systems.

The BH4 domain is an anti-apoptotic specific domain of the Bcl-2 protein. Solid phase peptide synthesis was used to produce large amount of the peptide for biophysical studies. A protocol has been established and optimized, guarantying the required purity for biophysical studies. In detail the purification by high performance liquid chromatography and the characterisation via mass spectroscopy are described. The secondary structure of BH4 changes significantly in the presence of lipid vesicles as observed by infrared spectroscopy and circular dichroism. The BH4 peptide aggregates at the membrane surface and inserts slightly into the hydrophobic part of the membrane. Using nuclear magnetic resonance (NMR) and calorimetry techniques, it could even be shown that the BH4 domain modifies the dynamic and organization of the liposomes which mimic a mitochondrial surface. The second study was on the first helix of the pro-apoptotic protein Bax. This sequence called Bax-α1 has the function to address the cytosolic Bax protein to the mitochondrial membrane upon activation. Once again a protocol has been established for the synthesis and purification of this peptide. The aim was to elucidate the key role of cardiolipin, a mitochondria-specific phospholipid, in the interaction of Bax-α1 with the mitochondrial membrane system. The NMR and circular dichroism studies showed that Bax-α1 interacts with the membrane models only if they contain the cardiolipin, producing a strong electrostatic lock effect which is located at the membrane surface.

Finally, a new NMR approach was developed which allows the investigation of the lipid response of isolated active mitochondria upon the presence of apoptotic stimuli. The goal was there to directly monitor lipid specific the occurring changes during these physiological activities.

Place, publisher, year, edition, pages
Umeå: Kemi, 2008. 58 p.
Apoptosis, BH4 peptide, Bax-α1 peptide, model membrane, cardiolipin, solid phase peptide synthesis, circular dichroism, solid-state nuclear magnetic resonance spectroscopy.
National Category
urn:nbn:se:umu:diva-1883 (URN)978-91-7264-676-6 (ISBN)
Public defence
2008-11-07, BiA201, Biologihuset, Umeå, 13:00 (English)
Available from: 2008-10-17 Created: 2008-10-17 Last updated: 2015-11-13Bibliographically approved

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