RESONANCE RAMAN CHARACTERIZATION OF NITRIC-OXIDE ADDUCTS OF CYTOCHROME-P450CAM - THE EFFECT OF SUBSTRATE STRUCTURE ON THE IRON LIGAND VIBRATIONS

Resonance Raman spectra of the nitric oxide adducts of ferric cytochrome P450cam are acquired with Soret excitation. The frequencies of the heme skeletal vibrations are found 3-10 cm-1 lower than those of horseradish peroxidase and myoglobin. The axial vibrations are located and assigned by isotopic substitution. The stretching, nu(Fe-NO), and bending, delta(Fe-N-O), modes are detected at 522 and 546 cm-1, respectively, for the title adduct in the presence of camphor. They shift to 520 and 533 cm-1 upon substitution by (NO)-N-15-O-16. The strong line at 522 cm-1 shifts to 528 cm-1 when the substrate is removed from the active site, whereas it appears at 524 cm-1 upon the addition of norcamphor. The isotopic sensitivity of these bands confirms their assignment as nu(Fe-NO). It is observed that only the larger substrates (camphor and adamantanone) give rise to the enhancement of the bending mode delta(Fe-N-O). As the substrate size increases, a band of medium intensity, which is tentatively assigned as nu(Fe-S), increases from 349 cm-1 (substrate free) to 359 cm-1 (adamantanone). On the basis of the consideration of both the electronic factors and the kinematic effect of distortion of the Fe-NO unit, it is suggested that the Fe-NO linkage adopts a linear structure in the absence of substrate but becomes slightly bent in the presence of substrates.