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Team:Copenhagen/Project/Water
From 2011.igem.org
water cleansing power
Background information:
A characterization of the cytochromes p450 is the main objective with this project. We will exploit the wide specificity of these cytochromes to target many different damaging agents continuously by hydroxylation, which can reduce the damaging activity of e.g. estrogen that it excerts when it binds to the estrogen receptors to often.
Our solution to that problem is to change the estrogen into a compound that has a low affinity for the estrogen receptors. 2-hydroxyestradiol with an affinity for the estrogen receptor at about 25% of estrogen is such a compound(1). The CYPs we'll explore are CYP1A1, CYP1A2, CYP2C9 and CYP3A4.
The human isoforms CYP1A2 has been proven to have a very high catalytic activity for hydroxylation of the number 2 carbon of estradiol in addition to a lower catalytic activity for the number 4 carbon(2). In addition it has been shown that CYP2C9 has a very specific catalytic activity this carbon as well(2). The mechanism that the CYP's use is shown below
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The project:
It is our intention to insert a plasmid into E. Coli containing a promoter that we control as well as the sequence for one of the CYP's. After that we will turn our bacteria into cytochrome p450 factories in order to create enough so that they can convert a lot of 17beta-estradiol into 2hydroxyestradiol. As a courtesy of Quo Data we have been granted a A-Yes-Assay kit, that allows us to test how many compounds in water that reacts with the estrogen receptors. Hopefully if our CYP's are effective, we will observe that the ammount of compounds that react with the receptors will decrease.
Ecological and economical prospects:
Another future aim with our project is to improve wastewater treatments and thus contribute to a cleaner and uncontaminated drinking water for everyone. With this scientific project, we will move towards this realization, but more research is needed. Our belief is that further research on the use of cytochrome p450 will give a valid and an inexpensive product for use in wastewater treatment plants, thus benefitting the environment and our planet.
References:
1. Schutze, N., Vollmer, G., Wunsche, W., Grote, A., Feit, B., and Knuppen, R. (1994) Binding of 2-hydroxyestradiol and 4-hydroxyestradiol to the estrogen receptor of MCF-7 cells in cytosolic extracts and in nuclei of intact cells, Exp Clin Endocrinol 102, 399-408.
2. Lee, A. J., Cai, M. X., Thomas, P. E., Conney, A. H., and Zhu, B. T. (2003) Characterization of the oxidative metabolites of 17beta-estradiol and estrone formed by 15 selectively expressed human cytochrome p450 isoforms, Endocrinology 144, 3382-3398.
