DISCRIMINATION BETWEEN CONFORMATIONAL STATES OF MITOCHONDRIAL CYTOCHROME P-450SCC BY SELECTIVE MODIFICATION WITH PYRIDOXAL 5-PHOSPHATE

Electrophoretically homogeneous cytochrome P-450scc preparation isolat ed by the standard method from adrenal cortex mitochondria comprises t wo protein forms differing in the accessibility of their amine groups to specific chemical modification with pyridoxal 5-phosphate. The prot ein form whose lysine amino groups are accessible to the modifier cons titutes about 60-70% of the preparation. Being covalently bound to pyr idoxal 5-phosphate, this protein form loses enzymatic activity and aff inity for adrenodoxin. This protein form can be separated by affinity chromatography on adrenodoxin-Sepharose. The cytochrome P-450scc form whose amino groups are not accessible to the modifier is retained on t he affinity matrix, and after elution from adrenodoxin-Sepharose has t he absorption spectrum typical of the high-spin protein with a spectra l homogeneity index A(392)/A(278) = 1.0. The enzymatic activity of the hemoprotein form whose lysine amino groups are inaccessible to the mo dification is identical to that of the initial unmodified protein.