Making hyperfine selection in Mims ENDOR independent of deadtime

A form of remote echo-detected ENDOR can be performed by adding an additional π pulse to the detection phase of a Mims three-pulse, stimulated-echo ENDOR experiment. The resulting four-pulse experiment, denoted refocused Mims (ReMims) ENDOR, retains the hyperfine selectivity of the Mims technique, but makes it possible to obtain arbitrarily short preparation intervals, τ1, independent of the spectrometer deadtime. As a result, without resonator redesign, it is possible to study a wider range of hyperfine values without distortions from “blind-spots”. The hyperfine selectivity of the ReMims ENDOR sequence is demonstrated on the 1H and 14N ENDOR spectra from a single crystal of Cu(glycine)2. Use of the ReMims protocol to augment the basic Mims and Davies ENDOR sequences is shown in two different types of situation: the sign of a 9 MHz coupled proton is assigned by the use of the implicit-TRIPLE effect in the proton ENDOR spectrum of nitrile hydratase; the true lineshape of the proton ENDOR spectrum of fluorometmyoglobin is determined without distortions by blindspots.