NMR structural and functional studies of the mithochondrial outer membrane protein VDAC by Thomas J. Malia.

Apoptosis is a mechanism of programmed cell death required by multicellular organisms during development and for tissue maintenance. Bcl-2 family proteins are central regulators of apoptosis and many of their primary roles are carried out in the outer mitochondrial membrane. Anti-apoptotic Bcl-xL has been found previously to interact with VDAC, an outer mitochondrial membrane protein responsible for metabolite trafficking. Here, the NMR-based investigation of VDAC and its interaction with Bcl-xL in detergent micelles is described. As an integral membrane protein VDAC presented a challenge for producing a folded form amenable to solution NMR studies. Methods developed for expression and purification of human VDAC for structural studies by NMR spectroscopy were developed. Optimal sample conditions were explored in a screen of various detergents and buffers. Suitable NMR sample conditions for VDAC were identified and allowed further characterization of VDAC and its interactions by NMR and other methods. Obtained through various means, evidence of a concentration dependent self-association of VDAC is also presented. Chemical cross-linking, analytical size exclusion chromatography, and NMR spectroscopic studies strongly support an equilibrium model for LDAO-purified VDAC between monomer and trimer.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemistry.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2006

Author dc:creator

• Malia, Thomas J., 1977-

Advisor dc:contributor.advisor

• Gerhard Wagner.

• Chemistry.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/37889

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/37889