Solid-state NMR studies of enzyme mechanism and membrane protein structure
NMR Structural Studies of Membrane Proteins: Most membrane-bound proteins are structurally uncharacterized at present; solid state NMR methods promise to offer important information for these systems. Recently we discovered that NMR spectra of uniformly labeled solid state proteins are well-resolved and may provide the basis for structural and functional studies. Many small proteins, including BPTI and ubiquitin, and several intrinsic membrane proteins have been studied using high-field state-of-the-art solid state NMR equipment. Torsional angles and tertiary contacts are characterized through existing dipolar methods.
Enzymes, Hydrogen Bonding Geometry, and Dynamics: For most enzymes and drug targets, ligand binding is associated with the motion of a flexible loop or domain and the restructuring of hydrogen bonds and other contacts. The characteristic timescales of an active-site flexible loop in TIM is under investigation. Similarly, metal-substrate geometry as well as conformational exchange rates are studied for metalloenzymes, such as the important drug target cytochrome P450. NMR measurements in the active sites of enzymes give insight into catalytic mechanism, drug binding modes, and dynamics.
NMR Methods Development, Enhanced Signals, and Alignment Protocols: NMR signals associated with the photosynthetic apparatus appear with intensities enhanced 300 to 1,000 times, relative to control values. We have proposed a mechanism for this remarkable effect and achieved quantitative agreement. We have demonstrated alignment of liquids in the presence of large AC electric fields, as detected by NMR spectroscopy. This experiment benefits simultaneously from the advantages of NMR for studying both solids and liquids. Ongoing design and optimization of NMR hardware for these and other applications is a central activity in our research group.
- “Characterization of Slow Conformational Dynamics in Solids: Dipolar CODEX” W. Li, A. E. McDermott, J. Biomol. NMR 45(1-2), 227-232 (2009)
- “Monitoring Conformational Dynamics with Solid-State R1roh Experiments” C. M. Quinn, A. E. McDermott, J. Biomol. NMR 45(1-2), 5-8 (2009)
- “Structure and Dynamics of Membrane Proteins by Magic Angle Spinning Solid-State NMR” A. McDermott, Ann. Rev. Biophys 38, 385-403 (2009)
- “Solid-State NMR on a Type III Antifreeze Protein in the Presence of Ice” A. B. Siemer, A. E. McDermott, J. Am. Chem. Soc. (2008)
- “Conformational Dynamics of an Intact Virus: Order Parameters for the Coat Protein of Pf1 Bacteriophage” J. L. Loriaeu, L. A. Day, A. E. McDermott, Proc. Nat. Ac. Sci. 105(30), 10366-10371 (2008)
- “Partial Site-Specific Assignment of a Uniformly 13C, 15N Enriched Membrane Protein, Light-Harvesting Complex 1 (LH1), by Solid State NMR” L. Huang, A. E. McDermott, Biochim. Biophys. Acta 1777(9), 1098-108 (2008)
- “Solid-State NMR Study and Assignments of the KcsA Potassium Ion Channel of S. Lividans” K. Varga, L. Tian, A. E. McDermott, Biochim. Biophys. Acta 1774 (12), 1604-1613 (2007)
- “Solid State NMR: New Tools for Insight into Enzyme Function” A. E. McDermott, T. Polenova, Curr. Opin. Struct. Biol. 17 (5), 617-622 (2007)
- “Assignment of Congested NMR Spectra: Carbonyl Backbone Enrichment Via the Entner-Doudoroff Pathway” A. Goldbourt, L. D. Day, A. E. McDermott, J. Magn. Reson. 189, 157-165 (2007)
- “Cytochrome P450 BM-3 in Complex with Its Substrate: Temperature-Dependent Spin State Equilibria in the Oxidized and Reduced States” T. Jovanovic, M. Harris, A. E. McDermott, App. Magn. Reson. 31 (3-4), 411-429 (2007)