Construction of Environmental Biremediation Enzymes Whose Catalysis is Regulated by Light and Molecular Switches

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Cytochrome P450s activate carcinogenetic activities and toxicities of various environmental chemicals including benzpyrene, aflatoxin, haloethanes. However, cytochrome P450s are also involved in metabolism and degradation of various environmental chemicals. Therefore, it is possible that we can construct efficient and useful environmental bioremediation enzymes if the degradation properties of cytochrome P450s can be used. A physiologically important gas, nitric oxide (NO), is synthesized by nitric oxide synthase whose active site structure is the sameas those of cytochrome P450s. On the other hand, there are many sensor enzymes and sensor proteins, where electron introduction, light illumination, bindings of calcium ion, CO, NO, molecule oxygen and various carcinogens induced certain protein conformational changes. The changes became signals that transmit to catalytic domain and switch the catalysis on/off. In the present study, we examined electron transfer process and protein-protein interactions of a calcium sensor protein, neuronal NO synthase. Then, we constructed chimeric enzymes composed of the calcium sensor domain of NO synthase and the have-bound active site of cytochrome P450. We examined efficiencies of electron transfer and catalysis of the generated calcium sensor chimeric enzymes with the an of a novel molecular switching enzyme. It was found that the chimeric enzyme functions as a calcium sensor enzyme. Furthermore, we examined structure and function relationships of a new redox sensor enzyme, Ec DOS. It was found that Ec DOS is also CO and NO sensor enzyme, because the catalysis was strongly inhibited by CO and NO. As the last step of this project study, we characterized potential heme-sensor enzyme, SOUL. In summary, the study provided the possibility to construct novel sensor enzymes with efficient biodegradation properties of environmental chemicals.