Seminar Title: "Magnetic Resonance Studies of Membrane Proteins"
Gary Lorigan, Professor of Chemistry and Biochemistry
Abstract: CW and pulsed Electron Paramagnetic Resonance (EPR) spectroscopic techniques coupled with site-directed spin-labeling (SDSL) can provide important structural information on complicated biological systems such as membrane proteins. Strategically placed spin-labels alter relaxation times of NMR active nuclei and yield pertinent structural information. EPR techniques such as Double Electron-Electron Resonance (DEER) and Electron Spin Echo Envelope Modulation (ESEEM) are powerful structural biology tools. The DEER technique can be used to measure distances between 2 spin labels from 20 to 70 Å. However, the application of DEER spectroscopy to study membrane proteins can be difficult due to short phase memory times (Tm) and weak DEER modulation in more biologically relevant proteoliposomes when compared to water soluble proteins or membrane proteins in detergent micelles. The combination of these factors often leads to broad distance distributions, poor signal to noise, and limitations in the determination of longer distances. The short phase memory times are typically due to uneven distributions of spin-labeled protein within the lipid bilayer, which creates local inhomogeneous pockets of high spin concentrations. Approaches to overcome these limitations and improve the quality of DEER measurements for membrane proteins will be discussed: lipodisq nanoparticles, bi-functional spin labels (BSL), and Q-band pulsed EPR spectroscopy. ESEEM data will be shown to probe the secondary structure of membrane proteins. CW-EPR spectra of spin-labeled membrane proteins will be used to investigate dynamics and the immersion depth in a lipid bilayer.
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