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Tracking lipid interactions in intact mitochondria under oxidative stress by ex vivo solid state 31P NMR spectroscopy
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).ORCID iD: 0000-0002-0546-7721
Umeå University, Faculty of Science and Technology, Department of Plant Physiology. Umeå University, Faculty of Science and Technology, Umeå Plant Science Centre (UPSC).ORCID iD: 0000-0001-5900-7395
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(English)In: Nature Methods, ISSN 1548-7091, E-ISSN 1548-7105Article in journal (Refereed) Submitted
Identifiers
URN: urn:nbn:se:umu:diva-3554OAI: oai:DiVA.org:umu-3554DiVA: diva2:142308
Available from: 2008-10-17 Created: 2008-10-17 Last updated: 2017-12-14Bibliographically 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.
Keyword
Apoptosis, BH4 peptide, Bax-α1 peptide, model membrane, cardiolipin, solid phase peptide synthesis, circular dichroism, solid-state nuclear magnetic resonance spectroscopy.
National Category
Biophysics
Identifiers
urn:nbn:se:umu:diva-1883 (URN)978-91-7264-676-6 (ISBN)
Public defence
2008-11-07, BiA201, Biologihuset, Umeå, 13:00 (English)
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Available from: 2008-10-17 Created: 2008-10-17 Last updated: 2015-11-13Bibliographically approved

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Sani, Marc-AntoineKeech, OlivierGardeström, PerGröbner, Gerhard
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