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Purification and Characterization of Overproduced E. coli Laccase  

Hong, June-Hyuk (Department of Biotechnology, Chung-Ang University)
Kim, Hyun-Jung (Department of Biotechnology, Chung-Ang University)
Kim, Woo-Yeon (Department of Biotechnology, Chung-Ang University)
Publication Information
Applied Biological Chemistry / v.50, no.2, 2007 , pp. 107-110 More about this Journal
Abstract
The putative laccase gene, yacK of Escherichia coli, K-12 is not expressed in lab culture conditions. The laccase gene was amplified by PCR and subcloned into pET28a vector. The laccase overproduced in E. coli harboring pET28a was purified by His-affinity column chromatography. The purified laccase, which has the apparent molecular weight of 55,000 on the SDS-polyacrylamide gel showed enzyme activity on the guaiacol solution and agar plate. Optimum temperature and pH were around 65$^{\circ}C$ and 5.0, respectively.
Keywords
E. coli laccase expression; His-affinity column purification;
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1 Alexandre, G. and Zhulin, L. B. (2000) Laccases are widespread in bacteria. Trends Biotechnol. 18, 41-42   DOI   ScienceOn
2 Andersen, S. O. (1985) Sclerotization and tanning of the cuticle, In: G. P. Kerkut and L. I. Gilbert (eds). Comparative Insect Physiology, Biochemistry, and Pharmacology, Vol. 3. Pergamon Press, New York, pp 59-74
3 Peter, M. G. and Wollenberg, U. W. (1997) Phenol-oxidizing enzymes: Mechanisms and applications in biosensors. Frontiers in Biosensorics I, pp 63-82
4 Mayer, A. M. and Harel, E. (1979) Polyphenoloxidases in plant. Phytochemistry 18, 193-215   DOI   ScienceOn
5 Liu, W., Chao, Y., Liu, S., Bao, H. and Qian, S. (2004) Molecular cloning and characterization of a laccase gene from the basidiomycete Fome lignosus and expression in Pichia pastoris. Appl. Microbiol. Biotechnol. 63, 174-181
6 Andersen, S. O., Peter, M. G. and Roepstorff, P. (1996) Cuticular sclerotization in insects. Comp. Biochem. Physol. 113B, 689-705
7 Pelaez, F., Martinez, M. J. and Martinez, A. T. (1995) Screening of 68 species of basidiomycetes for enzymes involved in lignin degradation. Mycol. Res. 99, 37-42   DOI   ScienceOn
8 Doerge, D. R., Divi, R. L. and Churchwell, M. I. (1997) Identification of the Colored Guaiacol Oxidation Product Produced by Peroxidases. Anal. Biochem. 250, 10-17   DOI   ScienceOn
9 Mayer, A. M. (1987) Polyphenoloxidase in plants-recent progress. Phytochemistry 26, 11-20   DOI   ScienceOn
10 Dean, J. F. D. and Eriksson, K. E. L. (1994) Laccase and the deposition of lignin. Holzforschung 48, 21-33   DOI   ScienceOn
11 Kim, C. W., Lorenz, W., Hoopes, T. and Dean, J.F.D. (2001) Oxidation of phenolate siderophores by the Multicopper oxidase encoded by the Escherichia coli yacK gene. J. Bacteriol. 183, 4866-4875   DOI   ScienceOn
12 Raghukumar, C., Raghukumar, S., Chinnaraj, A., Chandramohan, D., Dsouza, T. M. and Reddy, C. A. (1994) Laccase and other lignocillulose modifying enzymes of marine fungi isolated from the coast of India. Bot. Marina 37, 515-523   DOI   ScienceOn
13 Solomon, E. I., Sundaram, U. M. and Machonkin, T. E. (1996) Multicopper oxidases and oxygenases. Chem. Rev. 96, 2563- 2605   DOI   ScienceOn
14 Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680-685   DOI   ScienceOn
15 Link, A., Robinson, J., K. and Church, G. M. (1997) Comparing the predicted and observed properties of proteins encoded in the genome of Escherichia coli K-12. Electrophoresis 18, 1259-1313   DOI   ScienceOn