Chimeric enzymes with novel catalytic combinations have been created for various purposes, especially to improve kinetic properties and catalytic performances of the parental enzymes. Herein, a bifunctional enzyme was constructed by fusing catalytic conserved region of CotA from Bacillus subtilis to glutathione S-transferase (GST) to produce a chimera exhibiting both laccase and GST activities. Enzyme assays revealed that the chimeric GST-CotA protein, which was expressed in Escherichia coli BL21 harboring recombinant pGEX-4T-2 cloning vector, preserved both parental catalytic properties. Furthermore, GST activity of GST-CotA was about 12% higher than that of the parental GST, suggesting improved enzyme activity. The proteomic profile analysis by SDS-PAGE technique confirmed that a 36 kDa-protein corresponding molecular weight of GST-CotA was efficiently produced by E. coli. Considering the versatility of laccase and GST, this chimeric enzyme could be potentially used for various biotechnological applications, particularly for detoxification and bioremediation of a broad range of environmental pollutants.