Purification and characterization of a thermostable laccase from the ascomycetes Cladosporium cladosporioides and its applications

Abstract

The fungus Cladosporium cladosporioides was isolated from a coal sample as a dye decolorizing microorganism. The maximum production of laccase from this fungus was found to be 4160U/l with 0.05mM CuSO4, 4% (w/v) glucose and lignin 0.04% (w/v). The laccase was purified from C. cladosporioides to homogeneity in a very active state using ammonium salt precipitation and gel permeation chromatography. Analysis with SDS–PAGE showed that the purified laccase was a monomeric protein of 71.2kDa, and the apparent molecular mass of this enzyme was 75.17kDa (m/z=75,174), which was accurately determined by MALDI/TOF-MS. The UV–vis absorbance and electron paramagnetic resonance spectra of the purified laccase showed the presence of type 1 and type 3 copper ions. The optimum pH and temperature of the purified laccase were 3.5 and 40–70°C, respectively, and the N-terminal amino acid sequence was determined to be IGPTGDMYIVNEDVS. The purified laccase was effective in the biotransformation of aromatic compounds as well as in the decolorization of various dyes.