Large-Volume Dielectric Resonators for mm-Wave Pulsed EPR

Pulse Electron Paramagnetic Resonance (EPR) spectroscopy is a well-developed indispensable tool for
solving complex problems in biochemistry and structural biology. Pulse EPR encompasses several established
methods such as double electron-electron resonance (DEER), relaxation-induced dipolar modulation
enhancement (RIDME), and double quantum coherence (DQC) spectroscopy that are uniquely suited for
obtaining long-range (up to 10 nm or longer) distances and distance distributions between spin labeled sites in
biomacromolecules. The distances between endogenous paramagnetic metal centers can also be determined.
Another particularly impactful application of pulse EPR is in studies of metalloenzymes, where such methods as
3-pulse electron spin echo envelope modulation (ESEEM) spectroscopy, electron nuclear double resonance
(ENDOR), hyperfine sublevel correlation spectroscopy (HYSCORE), and electron double resonance detected
NMR (EDNMR) spectroscopy have evolved into highly informative techniques for providing mechanistic details
on the function of many important metalloenzymes without the need of preparing protein crystals. However, for
all these pulse methods insufficient concentration sensitivity remains the main roadblock towards their broader
applicability in biochemical and biophysical research. The goal of this technology development project is to
dramatically - by at least one to two orders of magnitude - increase concentration sensitivity of pulse W-band
(94 GHz) EPR spectroscopy by developing large-volume, high-quality factor (high-Q), and high finesse
microwave resonators based on low-loss dielectric materials. So far this new pulse EPR technology has been
only demonstrated as a proof-of-principle and will require further substantial efforts including: (Aim 1) Design
and optimization of high volume / high quality factor resonators for pulse EPR for the best concentration
sensitivity and construction of the resonator prototypes; (Aim 2) Development of critical mm-wave
instrumentation to implement high volume / high sensitivity resonators for routine and advanced W-band pulse
EPR experiments; and (Aim 3) Validating the new technology using well studied and well characterized systems.
The latter will be done by carrying out 14N ESEEM/HYSCORE experiments with Fe center in myoglobin and di-
Mn complexes, evaluating 55Mn EDNMR sensitivity gains for a series of Mn ion complexes, including di-Mn
clusters as well as S3 state of the oxygen evolving complex of PSI, and applying the DEER method to resolve
distance and angular information for a series of rigid biradicals, transmembrane peptides and rhodopsin
oligomers. If successful, this project will provide an innovative technology solution to the paramount long-
standing issues of pulse EPR spectroscopy such as increasing both the excitation bandwidth and sensitivity at
higher EPR frequencies. This will lead to a broader applicability of pulse EPR methods across the fields of
biochemistry and structural biology.