Advanced paramagnetic resonance techniques, such as electron-nuclear double resonance (ENDOR) and electron spin-echo envelope modulation (ESEEM) spectroscopies are essential tools for determining metalloenzyme structure and function. However, we find that a complete and detailed investigation of a protein metal center requires the integration of complementary information from spectrometers with CW and pulsed capabilities that operate at multiple frequencies, including X and Q bands but also a still higher microwave frequency. The present proposal shows that W band (95 GHz) is optimal for this purpose. It thus seeks funds for the design and construction of a W-band, CW and pulsed, ENDOR and ESEEM spectrometer and for the development of the strategies and protocols that optimize its use in the context of a multi-frequency, multi-technique approach to the determination of metalloenzyme active-site structure and function.
