High Field DNP and EPR in Biological Systems

Massachusetts Institute Of Technology, Cambridge MA

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Abstract

DESCRIPTION (provided by applicant): This proposal is focused on the development of high frequency dynamic nuclear polarization (DNP) nuclear magnetic resonance (NMR) to enhance sensitivity in NMR based structural biology experiments. 1 CW DNP at 527 GHz/800 MHz: We plan to complete development of a 527 GHz/800 MHz DNP spectrometer. It will involve (1) fabrication of low temperature (LT), 4 channel MAS probes, (2) the cryogenics for MAS at 80 K, (3) corrugated waveguide, and (4) interfacing these components with a 527 GHz tunable gyrotron nearing completion. This will be the sole 527 GHz/800 MHz DNP instrument available to the research community in North America. 2 Applications to Peptides and Proteins: We plan to continue application of DNP to studies of proteins with the goal of optimizing the signal enhancements for 2H labeled proteins where we have recently observed ?=120-140, which is ~x4 larger than for 1H species. Experiments are planned to determine the structure of amyloid fibrils and the H+ conduction mechanism of bacteriorhodopsin (bR). 3 High B0 Cross Effect (CE) Polarizing Agents: We plan to develop new polarizing agents for CE DNP at 600-800 MHz. They will use larger e--e- dipole couplings, trityl-TEMPO biradicals, and narrow line radicals with different isotropic g-values that satisfy the cross effect matching condition. 4 Overhauser Effect (OE) DNP in Solids: We plan to develop narrow line radicals with large 1H hyperfine couplings that mediate OE DNP. The temperature, ?1S and ?0S dependence of these radicals will be studied. Present data indicates that OE enhancement scales with B0 so this could be the method of choice for high field DNP experiments. 5 Pulsed DNP Experiments: The specific aim is to move pulsed DNP to 140 GHz using our rebuilt EPR spectrometer, a 20 watt 140 GHz EIK (amplifier), a gyroamplifier under development, and a MAS probe with a TE011 resonator and 1.3 mm rotor. We will investigate NOVEL, the integrated solid effect (ISE), the dressed state solid effect (DSSE), and NRF-DNP as approaches for time domain polarization transfer. 6 Electron Decoupling: We have recently documented substantial signal losses (~50%) associated with the presence of DNP polarizing agents and MAS due to modulation of the e- 1H dipole coupling. We plan to recover these signals with electron decoupling similar to TPPM used for decoupling 1H-13C MAS spectra.