Purification and characterization of a new laccase from Shiraia sp.SUPER-H168

A new laccase from Shiraia sp.SUPER-H168 was purified by ion exchange column chromatography and gel permeation chromatography and the apparent molecular mass of this enzyme was 70.78 kDa, as determined by MALDI/TOF-MS. The optimum pH value of the purified laccase was 4, 6, 5.5 and 3 with 2,6-dimethoxyphenol (DMP), syringaldazine, guaiacol and 2,2'-azinobis-(3-ethylbenzthiazoline-6-sulphonate)(ABTS) as substrates, respectively. The optimum temperature of the purified laccase was 50 degrees C using DMP, syringaldazine and guaiacol as substrates, but 60 degrees C for ABTS. Inhibitors and metal ions of SDS, NaN3, Ag+ and Fe3+ showed inhibition on enzyme activity of 10.22%, 7.86%, 8.13% and 67.50%, respectively. Fe7+ completely inhibited the purified laccase. The K-cat/K-m values of the purified laccase toward DMP, ABTS guaiacol and syringaldazine were 3.99 x 10(6), 3.74 x 10(7), 8.01 x 10(4) and 2.35 x 10(7) mol(-1) LS-1, respectively. The N-terminal amino acid sequence of the purified laccase showed 36.4% similarity to Pleurotus ostrestus. Approximately 66% of the Acid Blue 129 (100 mg L-1) was decolorized by 2.5 U of the purified laccase after a 120 min incubation at 50 degrees C. Acid Red 1 (20 mg L-1) and Reactive Black 5 (50 mg L-1) were decolorized by the purified laccase after the addition of Acid Blue 129 (100 mg L-1). (C) 2012 Elsevier Ltd. All rights reserved.