Dipolar dephasing for structure determination in a paramagnetic environment

We demonstrate the efficacy of the REDOR-type sequences in determining dipolar coupling strength in a paramagnetic environment. Utilizing paramagnetic effects of enhanced relaxation rates and rapid electronic fluctuations in Cu(II)-(DL-Ala)2.H2O, the dipolar coupling for the methyl C–H that is 4.20 ​Å (methyl carbon) away from the Cu2+ ion, was estimated to be 8.8 ​± ​0.6 ​kHz. This coupling is scaled by a factor of ~0.3 in comparison to the rigid limit value of ~32 ​kHz, in line with partial averaging of the dipolar interaction by rotational motion of the methyl group. Limited variation in the scaling factor of the dipolar coupling strength at different temperatures is observed. The C–H internuclear distance derived from the size of the dipolar coupling is similar to that observed in the crystal structure. The errors in the dipolar coupling strength observed in the REDOR-type experiments are similar to those reported for diamagnetic systems. Increase in resolution due to the Fermi contact shifts, coupled with MAS frequencies of 30–35 ​kHz allowed to estimate the hyperfine coupling strengths for protons and carbons from the temperature dependence of the chemical shift and obtain a high resolution 1H–1H spin diffusion spectrum. This study shows the utility of REDOR-type sequences in obtaining reliable structural and dynamical information from a paramagnetic complex. We believe that this can help in studying the active site of paramagnetic metalloproteins at high resolution. © 2021 The Authors