Conformational change of cytochrome P450 1A2 induced by phospholipids and detergents

Recently, it was reported that the activity of rabbit P450 1A2 is markedly increased at elevated salt concentration (Yun, C-H., Song, M., Ahn, T., and Kim, H. (1996) J. Biol, Chem, 271, 31312-31316), The activity increase of P450 1A2 coincides with the raised alpha-helix content and decreased beta-sheet content, The presence of phospholipid magnified this effect. Here, possible structural change of rabbit P450 1A2 accompanying the phospholipid-induced increase in its enzyme activity was investigated by circular dichroism, fluorescence spectroscopy, and absorption spectroscopy, Studies with the reconstituted system supported by cumene hydroperoxide or NADPH showed that the P450 1A2 activities were found to be dependent on the head group and hydrocarbon chain length of phospholipid, Phosphatidylcholines having short hydrocarbon chains with a carbon number of 8-12 were very efficient for reconstitution of the P450-catalyzed reactions supported by both cumene hydroperoxide and NADPH, It was found that the phospholipid increased the alpha-helix content and lowered the beta-sheet content of P450, Intrinsic fluorescence intensity is also increased in the presence of phospholipid. The low spin iron configuration of P450 1A2 shifted toward the high spin configuration by most of the phospholipids in the endoplasmic reticulum, Some synthetic phospholipids having short hydrocarbon chains with a carbon number of 10-12 caused a shift in the spin equilibrium of P450 1A2 toward low spin, The effect of detergents on the activity and conformation of P450 1A2 was also studied, It was found that the addition of detergents to P450 1A2 solution increased the enzyme activity of P450 1A2. Detergents also increased the alpha-helix content and lowered the beta-sheet content of P450 1A2, Intrinsic fluorescence emissions also increased with the presence of detergents, Octyl glucoside and deoxycholate caused a shift toward high spin, On the other hand, cholate caused a shift toward low spin, It was found that the activity increase of rabbit P450 1A2 coincides with the conformational change including raised alpha-helix content. It is proposed that the interaction with the phospholipid molecules surrounding P450 1A2 in the endoplasmic reticulum is important for a functional conformation of P450 1A2 in a monooxygenase system including NADPH-P450 reductase.