Institution: | a Department of Biochemistry, Kagawa Medical School, Kagawa 761-07, Japan b Research Equipment Center, Kagawa Medical School, Kagawa 761-07, Japan c Department of Neuropsychiatry, Kagawa Medical School, Kagawa 761-07, Japan d Basic Research Laboratory, Himeji Institute of Technology, Himeji, Hyogo 671-22, Japan |
Abstract: | Cholesterol side-chain cleavage activities of cytochrome P-450ssc purified from bovine adrenocortical mitochondria were measured for various substrates, including cholesterol, 20S]-hydroxycholesterol, 22R]-hydroxycholesterol and 20R], R]-dihydroxycholesterol, in the reconstituted enzyme system at various Tween20 concentrations. The side-chain cleavage activity for cholesterol showed more than 10-fold enhancement upon addition of 0.1% Tween20, compared with that without the detergent. Addition of Tween20 did not cause any enhancement of the side-chain cleavage activities for 20S]-hydroxycholesterol and 22R]-hydroxycholesterol; rather, it resulted in an inhibition of the activities. The side-chain cleavage activity for 20R],22R]-dihydroxycholesterol showed a very high value even without the detergent. As the stimulatory effect of Tween20 was only specific for cholesterol, Tween20 seemed to enhance the rate of access of cholesterol to cytochrome P-450scc. These results are consistent with the suggestion that a transfer of substrate, cholesterol, in mitochondrial inner membrane, to the substrate-binding site of cytochrome P-450scc is the rate-limiting step in the cholesterol side-chain cleavage reaction. |