Electron spin-echo envelope modulation study of multicrystalline Cu2+- insulin: Effects of Cd2+ on the nuclear quadrupole interaction of the Cu2+-coordinated imidazole remote nitrogen

A comparison of electron spin-echo envelope modulation (ESEEM) spectra from multicrystalline Cu2+-insulin with and without additional Cd2+ show a dramatic change in the quadrupole coupling parameters of the remote nitrogens of the two histidine imidazoles that ligate to copper. Without Cd2+, the quadrupole parameters are like those observed in blue copper proteins and in copper substituted lactoferrin. With Cd2+ soaked into the Cu2+-insulin crystals, the quadrupole parameters are similar to those found in galactose oxidase. Theoretical simulations of ESEEM spectra guided by structure modeling suggest that these changes originate from differences in the hydrogen bonding environments of the imidazole remote nitrogen. In addition, a compilation of results from previous ESEEM studies of copper proteins reveals that the asymmetry parameter, η, may be an indicator of type of hydrogen bond the imidazole remote nitrogen makes. When η ≥ 0.9, the nitrogen hydrogen bonds to water, whereas when η < 0.9, the nitrogen hydrogen bonds to the protein.