Recombinant laccase production from Bacillus licheniformis O12: Characterization and its application for dye decolorization


Kesebir A. O. , KILIÇ D. , ŞİŞECİOĞLU M. , ADIGÜZEL A. , KÜFREVİOĞLU Ö. İ.

BIOLOGIA, 2021 (Journal Indexed in SCI) identifier identifier

  • Publication Type: Article / Article
  • Volume:
  • Publication Date: 2021
  • Doi Number: 10.1007/s11756-021-00847-1
  • Title of Journal : BIOLOGIA
  • Keywords: Bacillus licheniformis, Recombinant laccase, Kinetic parameters, Biochemical characterization, Decolorization, MOLECULAR-CLONING, PURIFICATION, ABILITY, FUNGAL, OXIDASE, ENZYME

Abstract

This study aimed to produce a novel, highly thermostable laccase from Bacillus licheniformis O12 bacterial DNA using a recombinant method and investigate the effects of some dyes found in textile wastewaters on decolorization. A putative laccase gene (CotA) from B. licheniformis O12 was cloned and expressed as a fusion protein in Escherichia coli BL21 (DE3) cells. The recombinant laccase was purified using the Ni-NTA affinity chromatography method. Accordingly, 411.76-fold purification was achieved with 0.014 U/mg specific activity, and the molecular mass of the enzyme was calculated as 66 kDa. The optimum temperature and pH values of the laccase were determined as 92 degrees C and pH 5.0, respectively. It was also found to be quite stable for 75 min under acidic and alkaline conditions. The activity was measured for 12 h at 60 degrees C and 92 degrees C. At 92 degrees C, it was observed that the activity halved after 12 h. The highest value at 60 degrees C was obtained at 9 h. While an activity decrease of approximately 10 % was observed in the first three hours, a slow increase was detected afterward. These results proved that the obtained laccase was highly resistant to pH and temperature. The recombinant laccase was significantly activated by Al3+, Cd2+, Cr2+, Cu2+, Fe2+, Hg2+, Pb2+, and was inhibited in the presence of organic solvents, surfactants, and laccase inhibitors. Finally, the effects of some textile dyes on decolorization were investigated using the fused recombinant laccase and found to be generally effective. The highest decolorization of the laccase treated with dye for 2 h was 51.2 % in acid black 1, while the lowest decolorization was 1.9 % in the congo red.