• Journal of Microbiology and Biotechnology
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• v.20 no.4
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• pp.712-717
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• 2010

Cytochrome P450 enzymes (P450s) are involved in the synthesis of a wide variety of valuable products and in the degradation of numerous toxic compounds. The P450 BM3 (CYP102A1) from Bacillus megaterium was the first P450 discovered to be fused to its redox partner, a mammalian-like diflavin reductase. Here, we report the development of a whole-cell biocatalyst using ice-nucleation protein (Inp) from Pseudomonas syringae to display a hemeand diflavin-containing oxidoreductase, P450 BM3 (a single, 119-kDa polypeptide with domains of both an oxygenase and a reductase) on the surface of Escherichia coli. The surface localization and functionality of the fusion protein containing P450 BM3 were verified by flow cytometry and measurement of enzymatic activities. The results of this study comprise the first report of microbial cell-surface display of a heme- and diflavin-containing enzyme. This system should allow us to select and develop oxidoreductases containing heme and/or flavins into practically useful whole-cell biocatalysts for extensive biotechnological applications, including selective synthesis of new chemicals and pharmaceuticals, bioconversion, bioremediation, live vaccine development, and biochip development.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.11a
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• pp.161-164
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• 1996

• Biomolecules & Therapeutics
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• v.20 no.6
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• pp.562-568
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• 2012

Cytochrome P450 BM3 (CYP102A1) from Bacillus megaterium is a self-sufficient monooxygenase that consists of a heme domain and FAD/FMN-containing reductase domain (BMR). In this report, the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) by BMR was evaluated as a method for monitoring BMR activity. The electron transfer proceeds from NADPH to BMR and then to BMR substrates, MTT and CTC. MTT and CTC are monotetrazolium salts that form formazans upon reduction. The reduction of MTT and CTC followed classical Michaelis-Menten kinetics ($k_{cat}=4120\;min^{-1}$, $K_m=77{\mu}M$ for MTT and $k_{cat}=6580\;min^{-1}$, $K_m=51{\mu}M$ for CTC). Our continuous assay using MTT and CTC allows the simple, rapid measurement of BMR activity. The BMR was able to metabolize mitomycin C and doxorubicin, which are anticancer drug substrates for CPR, producing the same metabolites as those produced by CPR. Moreover, the BMR was able to interact with CYP1A2 and transfer electrons to promote the oxidation reactions of substrates by CYP1A2 and CYP2E1 in humans. The results of this study suggest the possibility of the utilization of BMR as a surrogate for mammalian CPR.

• Journal of Biomedical Engineering Research
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• v.19 no.1
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• pp.81-90
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• 1998

Electromagnetic waves may induce various effects on nervous tissues either by thermal or non-thermal mechanisms. This paper intoduces a method to evalute the non-thermal effect to central nervous system by measuring the EEGs of the rabbits treated by nimodipine before exposed to weak microwave field. 20 rabbits were divided into 2 groups and their EEGs were measured after their head section were exposed to 2,450 MHz microwave with the power density of 10 dBm and 20 dBm respectively for 10 minutes and compared with those of the 3rd group of 10 rabbits which were not exposed. The 4th group of 10 rabbits were intravenously given with nimodipine before exposed to 20 dBm field to determine whether this drug would reverse the EEGs changes induced by weak microwave irradiation. As field poser exceeded 20 dBm although no significant physiological changes were observed, total induced EEGs power was remarkably decreased suggesting the presence of CNS activation. Using Fourier analysis on the EEGs signal it was found that remarkable decrease in delta band and increase in the alpha and beta bands in a significant manner(P<0.05) compared to control group. The changes were, however, not reversed by nimodipine-treatment. The effects may be pure thermal in nature because no significant change has been observed in nimodipine treated rabbits.