Time-resolved Fourier-transform infrared studies of the cytochrome P-450cam carbonmonoxide complex bound with (1R)-camphor and (1S)-camphor substrate.

The CO-binding reaction of cytochrome P-450cam bound with (1R)-camphor and (1S)-camphor are compared in the temperature region of 210-260 K using time-resolved Fourier-transform infrared spectroscopy with the CO stretch vibration as spectroscopic probe. For (1S)-camphor as substrate the association of CO is slowed down by a factor of 2, while the dissociation is accelerated by a factor of 3. The CO complex for the (1S)-camphor-bound P-450 is less stabilized (deltaG=-22 kJ/mol) compared to the natural substrate (1R)-camphor (deltaG=-30 kJ/mol). The data are interpreted by a smaller change of the mobility of the (1S)-camphor due to CO binding as compared to (1R)-camphor, which would indicate a higher mobility of (1S)-camphor already in the CO free reduced form of P-450cam. The higher mobility of (1S)-camphor in the heme pocket might explain the increased uncoupling rate (hydrogen peroxide formation) of 11% [Maryniak et al. (1993) Tetrahedron 49, 9373-9384] during the P-450cam catalyzed hydroxylation compared to 3% for the conversion of (1R)-camphor.