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Investigations of the biocatalytic activity of human P450 2D6Zhao, Jin, 1975- January 2007 (has links)
The cytochrome P450 enzymes (CYPs) are very attractive biocatalysts because of their ability to regio- and stereo-selectively catalyze the insertion of a single atom of molecular oxygen into inactivated C-H bonds. There are many drawbacks, however, limiting the use of these enzymes in organic synthesis, including the need for expensive cofactors, low stability, and low tolerance to organic solvents. The goal of this thesis was to overcome some of these drawbacks for human CYP2D6. This isoform was selected because of its broad substrate promiscuity and high importance in drug metabolism. / We have tested inexpensive chemicals to replace the natural cofactors of CYP2D6, NADPH and cytochrome P450 reductase (CPR). The results showed that cumene hydroperoxide and tert-butyl hydroperoxide can successfully substitute CPR and NAD(P)H with retained regio- and stereo-selectivity. Moreover, with these surrogates, product formation and initial rates are increased by as much as two fold compared to the use of the natural cofactors. / It is widely accepted that even small proportions of organic solvents in the buffer can deactivate most enzymes including P450s. Our studies on the biocatalysis of CYP2D6 in organic solvent/buffer emulsions showed that under the optimized conditions, as much as 76% of the enzyme activity was retained. Product formation in biphasic solvent systems is comparable whether the natural redox partner and cofactor are used, or a surrogate. In addition, a correlation was observed between the log P and the suitability of a solvent for enzymatic activity, with higher log P resulting in higher enzymatic activity. These results were obtained with dextromethorphan (DXM), a water soluble substrate. A very hydrophobic substrate, 7-benzyloxy-4-N,N-diethylaminomethyl-coumarin (BDAC), was also tested successfully to demonstrate the utility of this method. / Lyophilization is usually required to remove water before using enzymes in nearly anhydrous solvents. This physical process is harmful to P450 enzyme activity. We therefore tested numerous sugars as lyoprotectant during lyophilization. Addition of trehalose or sucrose before lyophilization allowed the retention of 80% of the CYP2D6 activity, compared to 40% remaining activity in its absence. CYP2D6 co-lyophilized with trehalose was next tested in selected hydrophobic organic solvents in the absence of water. The enzymatic activity was found to strongly depend on the hydrophobicity of the solvent. Interestingly, the enzyme showed higher catalytic ability in n-decane or n-dodecane than in the standard buffer. This was unexpected considering that the activity of most enzymes was reported to decrease to 10% or less in nearly anhydrous organic solvents. / The last objective of this thesis was to improve the stability and/or activity of CYP2D6. Use of DNA self-assemblies to encapsulate P450 enzymes was envisaged to potentially increase their stability. Indeed, DNA assemblies have many advantages compared to traditional solid supports reported for enzymes. Our preliminary results showed that CYP2D6 templated the formation of cyclic DNA dimeric and tetrameric over polymeric self-assemblies. Characterization of the CYP2D6 activity in the presence of the DNA self-assemblies revealed no loss of activity or stability.
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Investigations of the biocatalytic activity of human P450 2D6Zhao, Jin, 1975- January 2007 (has links)
No description available.
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The reaction of alternate oxidants with cytochrome P450BM3 and mutants generates spectrally detectable, high valent iron intermediatesThompson, Jonathan Ira. January 1900 (has links) (PDF)
Thesis (M.S.)--University of North Carolina at Greensboro, 2006. / Title from PDF title page screen. Advisor: Gregory Raner; submitted to the Dept. of Chemistry. Includes bibliographical references (p. 51-52).
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The effect of cyclophosphamide administration on the activity and relative content of hepatic CYP2D1 in the rat /Laslett, Tiffany Jane. Unknown Date (has links)
Thesis (MAppSc in Pharmacy)--University of South Australia, 1996
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Studies on the regulation of the barbiturate-inducible cytochrome P450 genes CYP2H1 and CYP2H2 : a thesis submitted for the degree of Doctor of Philosophy at the University of Adelaide / Benjamin Paul Davidson.Davidson, Benjamin Paul January 2000 (has links)
Bibliography: leaves 103-129. / x, 129, [63] leaves, [12] leaves of plates : ill. (chiefly col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / The study isolates a 920 bp proximal promoter segment of the CYP2H2 gene from a chicken genomic clone. / Thesis (Ph.D.)--University of Adelaide, Dept. of Biochemistry, 2000?
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Characterization of the chicken phenobarbital inducible P450 gene family /Elferink, Lisa Anne. January 1987 (has links) (PDF)
Thesis (Ph. D.)--University of Adelaide, 1977. / Includes bibliographical references.
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CYP2C19 catalyzed drug metabolism in different populations /Herrlin, Karin, January 1900 (has links)
Diss. (sammanfattning) Stockholm : Karol. inst., 2001. / Härtill 4 uppsatser.
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Regulation of the expression of phenobarbital-inducible P450 genes /Hansen, Antony James. January 1989 (has links) (PDF)
Thesis (Ph. D.)--University of Adelaide, Dept. of Biochemistry, 1990. / Includes bibliographical references.
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Studies on the regulation of the barbiturate-inducible cytochrome P450 genes CYP2H1 and CYP2H2 : a thesis submitted for the degree of Doctor of Philosophy at the University of Adelaide /Davidson, Benjamin Paul. January 2000 (has links) (PDF)
Thesis (Ph.D.)--University of Adelaide, Dept. of Biochemistry, 2000? / Bibliography: leaves 103-129.
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Lipid aldehydes are substrates and transcriptional regulators of cytochromes P450 /Amunom, Immaculate. January 2008 (has links) (PDF)
Thesis (Ph. D.)--University of Louisville, 2008. / Department of Biochemistry and Molecular Biology. Vita. "May 2008." Includes bibliographical references (leaves 189-200). Also in PDF.
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