Cytochrome P-450sccA Review of the Specificity and Properties of the Cholesterol Binding Site

Abstract

Cytochrome P-450_scc is unusual among members of this class of enzymes in showing a high degree of substrate specificity. Features of the cholesterol structure which are particularly important for binding include the 3$sZ-hydroxyl, the $DL 5-ring configuration, and the side-chain organization in the 20–22 region. Regarding the ring system, binding appears to require planarity and limited size at the 4–5–6 carbons (the A-B ring juncture). In the region of the 3$sZ-hydroxyl, a “cleft” in the binding site extends about 4 Å beyond the hydroxyl and can accommodate two additional ether-linked carbons. Evidence indicates that an enzyme residue hydrogen-bonds to the oxygen of the 3$sZ hydroxyl, providing much of the energy for the initial enzyme-substrate interaction. The cytochrome shows less specificity for the side-chain structure, except in the region of carbons 20–22 where hydroxylation/side-chain cleavage takes place. The binding cleft for the side-chain is limited to approximately the length of the isocaproic group but can accommodate structural variations beyond the 22-position. Evidence indicates that the region near the 20–22 bond is more limited in size, and that an amino acid residue near the heme iron binds strongly and stereospecifically to the 22R-hydoxyl of the cleavage intermediates, 22R-hydroxycholesterol and 20α,22R-dihydroxycholesterol. The 22R-hydrogen of cholesterol is very close to the heme iron (approximately 3 Å), while the 22S-hydrogen is slightly further (about 4 Å). The size and bonding properties of the steroid binding/active site suggest a mechanism which accounts for the stereospecificity and sequence of reactions catalyzed by cytochrome P-450_scc.