[Purification and properties of laccase from Basidiomycete].
Laccase produced by Basidiomycete was purified to electrophoretic homogeneity by the steps of ammonium sulfate precipitation, DEAE-cellulose and hydrophobic interaction column chromatography. Purification of about 318.4 fold was achieved with an overall yield of 18.6%. Its molecular weight was estimated to be about 60.3 kD by SDS-PAGE, and that of it was 55.94 kD by mass spectrum. The optimum temperature and pH of the enzyme activity were 65 degrees C and 2.2 - 2.8 respectively. The isoelectric point was 4.02 (room temperature). Its N-terminal sequence was AIGPVTDL. The carbohydrate content was 49.2% by the phenol-sulfuric acid method. Michaelis constant of the enzyme for ABTS was 17.5 micromol/L. The enzyme activity was stable under 45 degrees C and in the pH range of 3.0 - 9.5. The activity was enhanced by Cu2+, and was strongly inhibited by Fe2+. While Mn2+ and Ag+ had no effect on laccase activity. Dithiothreitol and sodium azide inhibited completely the activity. Trp was possible essential residue for enzyme activity.
Liu
Shuzhen
S
Institute of Microbiology, Chinese Academy of Sciences , Beijing 100080, China. szliu0252@sina.com
Qian
Shijun
S
D001487:Basidiomycota / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D042845:Laccase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D013696:Temperature
D042845:Laccase