| Popis: |
Earlier studies have shown that the reactions of cytochrome P450 with arylhydrazines yield aryl-iron complexes, and that oxidative migration of the aryl groups to the pyrrole nitrogens of the heme provides information on the active site topology. Comparison of cytochromes P450terp (CYP108), P450cam (CYP101), and P450BM-3 (CYP102) by this method suggests that the active site of P450terp is effectively more sterically restricted than those of the other two enzymes and is primarily open above pyrrole ring D of the heme group. This experimental model of the P450terp active site differs from that deduced by x-ray crystallography, which shows that pyrrole ring C is also relatively open. The results suggest that aryl shifts can be used to probe conformations of the active site other than that trapped in the crystal state. Identification of the product formed from alpha-terpineol by P450terp shows that the enzyme exclusively hydroxylates the most sterically accessible, allylically activated position. The enzyme also oxidizes substituted thioanisoles and styrenes unrelated to alpha-terpineol to the corresponding sulfoxides and epoxides. In the case of 4-methylthioanisole and 4-methylstyrene, methyl hydroxylation competes effectively with sulfoxidation and epoxidation in the reaction catalyzed by P450terp but not those catalyzed by P450BM-3 or P450cam. Comparison of the stereoselectively of thioanisole sulfoxidation and styrene epoxidation by P450terp, P450cam, and P450BM-3 shows that P450terp is the most, and P450BM-3 the least, stereospecific. The stereospecificity of thioanisole sulfoxidation by P450terp depends on the electronic nature of the para-substituent and rises from an (R):(S) ratio of 20:80 for p-MeO to a value of < 01:99 for p-CN. The (R):(S) ratio for the epoxides produced by P450terp is approximately 90:10 for the two substituents investigated. Cytochromes P450cam and P450BM-3 are much less stereoselective. A model is suggested by the stereochemical and topological data for the binding of substrates in P450terp. |
