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Title:Solid-state NMR studies of membrane proteins and membrane protein complexes
Author(s):Sperling, Lindsay J.
Director of Research:Rienstra, Chad M.
Doctoral Committee Chair(s):Rienstra, Chad M.
Doctoral Committee Member(s):Oldfield, Eric; Gruebele, Martin; Nair, Satish K.
Department / Program:Chemistry
Discipline:Chemistry
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):Cytochrome bo3 oxidase
DsbA/DsbB
Magic-angle Spinning
Membrane Proteins
Solid-state nuclear magnetic resonance (NMR)
Abstract:Membrane proteins help control nearly every process in the cell, which is why approximately 50% of pharmaceuticals currently on the market target membrane proteins. Knowledge of structure-function relationships of these proteins could be leveraged to produce more efficient drugs. However, the traits that make membrane proteins so interesting also make them difficult targets for traditional structure elucidation techniques. X-ray crystallography relies on the use of single crystals, production of which are elusive for membrane proteins due to their inherent dynamic loops and stretches of hydrophobic residues, which contribute to aggregation and/or loss of function without the presence of a lipid environment. Solution NMR experiences difficulty dealing with slow molecular tumbling due to the large sizes of membrane proteins. Conversely, solid-state NMR (SSNMR) has no inherent size limitation and does not require the use of crystals, which presents SSNMR with the unique capability to study membrane proteins in native environments at atomic-resolution. However, this technique is still a relatively new tool for solving structures of biomolecules. Here, we begin to develop strategies for solid-state NMR de novo structure determination. We provide a “divide-and-conquer” investigation of an E. coli 41 kDa membrane protein complex, DsbA/DsbB. We begin by completing chemical shift assignments, the first step in structure determination in NMR studies, of the 21 kDa protein DsbA to optimize sensitivity and resolution of data collection and analysis of large systems. We then use this study to drive forward structural examination of the disulfide bond forming system DsbA/DsbB. Finally, SSNMR techniques are used to study a 144 kDa cytochrome bo3 ubiquinol oxidase demonstrating the power of this technique to investigate large membrane complexes in native environments.
Issue Date:2011-08-26
URI:http://hdl.handle.net/2142/26275
Rights Information:Copyright 2011 Lindsay J. Sperling
Date Available in IDEALS:2011-08-26
2013-08-27
Date Deposited:2011-08


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