Two- and four-pulse ESEEM studies of the heme binding center of a low-spin ferriheme protein: the importance of a multi-frequency approach

The histamine complex of nitrophorin1 (NP1-Km) was studied by ESEEM at X- and S-bands. The proton sum combination feature in the X-band ESEEM spectra was well resolved at the low-field g-value(g(Z)), but unresolved at the high-field g-value (g(X)). Decreasing the microwave frequency down to S-band allowed us to obtain well-resolved ESEEM spectra at the g(X) position and to determine the orientation of g(X) to be 60 degrees-90 degrees with respect to the plane of the axial ligands. (C) 1999 Elsevier Science B.V. All rights reserved.