Mycosystema. 2007, 26(4): 539-548.
Laccase high-yielding strain SAH-12 of Trametes gallica was obtained by UV mutagenesis. Three isoenzymes of laccase (viz. Lac1, Lac2 and Lac3) were obtained in the culture solution of SAH-12, among which Lac1 of electrophoretic homogeneity was purified by the steps of ammonium sulfate precipitation, dialysis and Sephadex G-75 chromatography. Purification of about 6.54 fold was achieved with an overall yield of 59.7%. The molecular weight of purified Lac1 was estimated to be about 61.5kDa by SDS-PAGE. It was a glycoprotein with carbohydrate content of 11.6%. The isoelectric point was 4.40 (room temperature). The optimum temperature and pH of the Lac1 activity were 60℃ and 2.6, respectively in catalytic reaction of oxidizing ABTS. Michaelis constant of the enzyme for ABTS was 25.0μmol/L. The enzyme activity was stable under 40℃ and pH4.0, and within pH range of 1.5~5.0 under 28℃. The activity of Lac1 was enhanced by Cu2+, whereas it was inhibited by the metal ions Fe2+, Ag+, Hg2+ and Cr3+, and inhibitors DTT, SDS, EDTA and DMSO. The activities of Lac1 were completely inhibited by Fe2+ and DTT, while Mn2+ and Zn2+ did not strongly effect on laccase activity. Lac1 was not only able to decolorize some synthetic dyes efficiently, but also be able to digest polyphenol in apple juice. The polyphenol of apple juice decreased 40% by Lac1 at 40℃ for 5h.
