Structure and spin density of ferric low-spin heme complexes determined with high-resolution ESEEM experiments at 35...

The wide use of the heme group by nature is a consequence of its unusual "electronic flexibility." Major changes in the electronic structure of this molecule can result from small perturbations in its environment. To understand the way the electronic distribution is dictated by the structure of the heme site, it is extremely important to have methods to reliably determine both of them. In this work we propose a way to obtain this information in ferric low-spin heme centers via the determination of g, A, and Q tensors of the coordinated nitrogens using electron spin echo envelope modulation experiments at Q-band microwave frequencies. The results for two bisimidazole heme model complexes, namely, PPIX(Im)(2) and CPIII(Im)(2), where PPIX is protoporphyrin IX, CPIII is coproporphyrin III, and Im is imidazole, selectively labeled with (15)N on the heme or imidazole nitrogens are presented. The planes of the axial ligands were found to be parallel and oriented approximately along one of the N-Fe-N directions of the slightly ruffled porphyrin ring (approximately 10 degrees ). The spin density was determined to reside in an iron d orbital perpendicular to the heme plane and oriented along the other porphyrin N-Fe-N direction, perpendicular to the axial imidazoles. The benefit of the method presented here lies in the use of Q-band microwave frequencies, which improves the orientation selection, results in no/fewer combination lines in the spectra, and allows separation of the contributions of hyperfine and quadrupole interactions due to the fulfillment of the exact cancellation condition at g ( Z ) and the possibility of performing hyperfine decoupling experiments at the g ( X ) observer position. These experimental advantages make the interpretation of the spectra straightforward, which results in precise and reliable determination of the structure and spin distribution.