Low frequency spectral density of ferrous heme: Perturbations induced by axial ligation and protein insertion
Flaviu Gruia 1, Xiong Ye 1, Dan Ionascu 1, Minoru Kubo 1 and Paul M. Champion 2*
1 Northeastern University
2 Northeastern Univ.
* To whom correspondence should be addressed. E-mail: champ{at}neu.edu.
Submitted on June 11, 2007
Revised on August 9, 2007
Accepted on 13 August 2007
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Abstract |
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Femtosecond coherence spectroscopy is used to probe low frequency (20-400cm-1) modes of the ferrous heme group in solution, with and without 2-methyl imidazole (2MeIm) as an axial ligand. The results are compared to heme proteins (CPO, P450cam, HRP, Mb) where insertion of the heme into the protein results in redistribution of the low frequency spectral density and in (~60%) longer damping times for the coherent signals. The major effect of imidazole ligation to the ferrous heme is the "softening" of the low frequency force constants by a factor of ~0.6±0.1. The functional consequences of imidazole ligation are assessed and it is found that the enthalpic CO rebinding barrier is increased significantly. When combined with the kinetic results, the force constant softening analysis indicates that the iron is displaced by only ~0.2Å from the heme plane in the absence of the imidazole ligand, while it is displaced by ~0.4Å when imidazole (histidine) is present. This suggests that binding of imidazole (histidine) as an axial ligand, and the concomitant softening of the force constants, leads to an anharmonic distortion of the heme group that has significant functional consequences.
Key Words:
Raman spectroscopy, femtosecond coherence spectroscopy, heme-CO kinetics, heme-imidazole
