A thermodynamic-kinetic analysis of the cytochrome P-450 heme pocket.
Spin state changes in the iron center of cytochrome P-450 during the catalytic cycle suggest alterations in the heme environment that insure proper substrate binding, an increase in redox potential, the formation of an active Fe-O complex, and the attack on the substrate. We used the spin state changes of the iron following physico-chemical perturbations, as an intrinsic probe of discrete changes around the heme, or of larger ones in the protein conformation. These environmental perturbations included temperature, solvent, substrate, and ionic environment. Aqueous and hydro-organic buffers provide complementary data and interpretations; the mixed solvent accommodates temperatures suitable for direct reaction rate measurements and amplified low to high spin transition. The results suggest that the group determining the heme spin state is influenced by the electrostatic potential created by several negative charges near the heme; the modulation of the spin state by various factors reflects the modulation of the electrostatic potential and of the internal paH value. Conformational changes of the whole protein are also indicated by the large entropy terms and their variation with experimental conditions.
D001419:Bacteria / Q000378:metabolism; D002151:Calorimetry; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme; D007700:Kinetics; D011487:Protein Conformation; D013696:Temperature; D013816:Thermodynamics
D006418:Heme; D003577:Cytochrome P-450 Enzyme System