Cytochrome P-450-dependent digitoxin 12β-hydroxylase from cell cultures of Digitalis lanata

AbstractA 12β-hydroxylase which catalyzes the hydroxylation of digitoxin and other cardenolides of the A-series to digoxin and the corresponding cardenolides of the C-series was isolated from suspension cultures of Digitalis lanata. The enzyme is located in the microsomal fraction and has a pH optimum at 7.5. The Km value for β-methyldigitoxin is 7.1 μM. O2 and NADPH2 are essential for the catalytic activity, with an optimum NADPH2 concentration of 1 mM. Carbon monoxide inhibits the enzyme, but this inhibition is reversible with 450 nm light. This would suggest that the enzyme is a cytochrome P-450-dependent mixed-function monooxygenase. p-Hydroxymercuribenzoate and CuSO4 are also inhibitory. KCN stimulates hydroxylation in vitro