Funds are requested to complete the purchase of a Bruker EleXsys 580 Q-band FT-EPR system at Vanderbilt University. This equipment will be utilized by five different established research groups and a number of their collaborators to make DEER measurements on a wide range of nitroxide spin-labeled proteins that are already being studied using site-directed spin-labeling. Recent advances in instrumentation and methodology have positioned time-domain EPR as an essential complement to cw-EPR for characterizing the structures and dynamics of spin-labeled proteins. In the ten NIH funded projects that constitute the core user group of this application, the requested equipment will be utilized to: 1) increase our capacity for measuring """"""""long"""""""" (>20 - 25 E) inter-probe distances in proteins by DEER;2) increase signal to noise for small samples including reconstituted membrane proteins;3) permit multifrequency analysis of DEER data;4) provide orientation selectivity for assessment of relative orientations between spin labels;and 5) provide the capability to make reliable cw-Q-band measurements. In each of the five applicant laboratories, the requested instrumentation will have an immediate and substantial impact by providing unique new measurement capabilities for full characterization of the structures and dynamics of wide range of soluble and membrane bound proteins that are of clear biomedical importance. The requested instrumentation will also be made available to other investigators at Vanderbilt and in the surrounding area as they develop new projects that involve spin-labeling. Equitable utilization of the equipment will be assured by an internal oversight committee and day-to-day operation will be handled by an experienced EPR spectroscopist.
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