Abstract
Objective
To investigate the conversion of carbazole into 2′-aminobiphenyl-2,3-diol using carbazole dioxygenase (CARDO) that is a multicomponent enzyme consisting of homotrimeric terminal oxygenases (CarAa), a ferredoxin (CarAc) and a ferredoxin reductase (CarAd) unit, encoded by the carAa, carAc and carAd genes, respectively.
Results
The enzyme subunits containing a GST tag were expressed independently in E. coli. The expressed proteins were purified by one-step immobilized affinity chromatography and three purified proteins could reconstitute the CARDO activity in vitro and showed activity against carbazole as well as against wide range of polyaromatic compounds.
Conclusion
This method provides an efficient way to obtain an active carbazole dioxygenase with high yield, high purity and with activity against a wide range of polyaromatic compounds.
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Acknowledgments
This work is supported by the research funding provided by Department of Biotechnology, Government of India (Project no BT/PR7574/BCE/8/1001/2013).
Supporting information
Supplementary Table 1—Sequence of the primers used for amplification of different subunits of CARDO (CarAa, CarAc and CarAd.
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Khan, S., Adhikari, D.K., Gupta, S. et al. High-level expression, purification and characterization of carbazole dioxygenase, a three components dioxygenase, of Pseudomonas GBS.5. Biotechnol Lett 37, 1945–1952 (2015). https://doi.org/10.1007/s10529-015-1876-3
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DOI: https://doi.org/10.1007/s10529-015-1876-3