title: Rapid spectral scan and stopped-flow studies of carbon monoxide binding to bovine adrenocortical cytochrome P-450scc.
abstract: Carbon monoxide binding with both cholesterol-free (low-spin) and cholesterol-bound (high-spin) reduced forms of purified cytochrome P-450scc has been investigated by rapid-scan and stopped-flow spectrometry. CO binding occurs within 150 ms at 25 degrees C for both forms of P-450scc, with a typical absorption maximum at 450 nm. Isosbestic points occur at the following wavelengths: between reduced-CO and reduced cholesterol-free P-450scc at 434 and 471 nm; between reduced-CO and reduced cholesterol-bound P-450scc at 433 and 469 nm. Both the 'on' (k1) and 'off' rate constants (k-1) are found to be independent of pH between pH 5 and 9. The mean values of k1 for cholesterol-free (1.8 +/- 0.2) X 10(5) M-1 X s-1) and cholesterol-bound [1.9 +/- 0.1) X 10(5) M-1 X s-1) P-450scc are almost identical, while the mean value of k-1 for the former [2.3 +/- 0.3) X 10 s-1) is about double that of the latter [1.2 +/- 0.1) X 10 s-1). This suggests the instability of the reduced-CO complex in the absence of cholesterol.
mesh_terms: D000302:Adrenal Cortex / Q000201:enzymology; D000818:Animals; D002248:Carbon Monoxide / Q000378:metabolism*; D002417:Cattle; D002784:Cholesterol / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008928:Mitochondria / Q000201:enzymology; D010084:Oxidation-Reduction; D011485:Protein Binding; D013057:Spectrum Analysis
chemical_list: D002248:Carbon Monoxide; D003577:Cytochrome P-450 Enzyme System; D002784:Cholesterol