This is a Shared Instrumentation Grant proposal for the acquisition of a modern commercial CW and FT-EPR Bruker EleXsys X-band spectrometer for use by a core group of six established Principal Investigators and one beginning investigator from Universities in the Research Triangle area of North Carolina. Currently, there is no such instrument in the Triangle area. The ongoing research projects involve applications of modern time-domain EPR methods including pulsed saturation recovery and HYSCORE to a wide range of biomedical problems and systems from hybrid nanoscale assemblies for membrane protein biochips to lipid transfer proteins, self-assembly of tRNA and drug-delivery assisted by nanoparticle-templated virions, and the structures of the enzymes'catalytic sites. Currently, all these projects are either pursued with available continuous wave X-band EPR instruments that are at least 20 years old or in collaborating with other groups that have Bruker EleXsys X-band spectrometer available. With this proposed major upgrade all seven core research projects will benefit from recent developments in instrumentation and methodology of time-domain EPR methods at X-band. Specifically, we will be able to 1) obtain long-range (to up to 60-70 angstrom) distance constrains for large biomolecular assemblies and partially folded structures;2) directly measure electronic relaxation rates of specifically labeled proteins, RNAs, and phospholipids in the presence and absence of paramagnetic relaxation agents in order to gain valuable structural information;and 3) obtain unique information on paramagnetic metal ion coordination upon substrate binding to catalytic enzymes (HYSCORE experiment).
The relevance of this project to public health will be in providing primarily NIH-funded investigators as well as other investigators from NCSU, UNC, and the Research Triangle area with an advanced FT- EPR spectrometer as these investigators develop new projects involving structure-function studies of proteins and RNAs. The new FT-EPR spectrometer will also enhance training of graduate students from the newly established NCSU RNA Biology/Chemistry -Biology Interface Training Program that involves ten NCSU Departments.
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|Li, Jikun; Wang, Ke; Smirnova, Tatyana I et al. (2013) Isoprenoid biosynthesis: ferraoxetane or allyl anion mechanism for IspH catalysis? Angew Chem Int Ed Engl 52:6522-5|
|Zhao, Qingbiao; Nellutla, Saritha; Son, Won-Joon et al. (2011) Ba4KFe3O9: a novel ferrite containing discrete 6-membered rings of corner-sharing FeO4 tetrahedra. Inorg Chem 50:10310-8|
|Wang, Weixue; Wang, Ke; Li, Jikun et al. (2011) An ENDOR and HYSCORE investigation of a reaction intermediate in IspG (GcpE) catalysis. J Am Chem Soc 133:8400-3|
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