Purification and characterization of an extracellular laccase from the edible mushroom Lentinula edodes, and decolorization of chemically different dyes.
A laccase (EC 1.10.3.2) was isolated from the culture filtrate of Lentinula edodes. The enzyme was purified to a homogeneous preparation using hydrophobic, anion-exchange, and size-exclusion chromatographies. SDS-PAGE analysis showed the purified laccase, Lcc 1, to be a monomeric protein of 72.2 kDa. The enzyme had an isoelectric point of around pH 3.0. The optimum pH for enzyme activity was around 4.0, and it was most active at 40 degrees C and stable up to 35 degrees C. The enzyme contained 23.8% carbohydrate and some copper atoms. The enzyme oxidized 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt, p-phenylendiamine, pyrogallol, guaiacol, 2,6-dimethoxyphenol, catechol, and ferulic acid, but not veratryl alcohol, tyrosine, and beta-(3,4-dihydroxyphenyl) alanine. The N-terminal amino acid sequence of Lcc 1 showed close homology to the N-terminal sequences determined for laccases from Phlebia radiata, Trametes villosa, and Trametes versicolor, but only low similarity was observed to a previously reported laccase from L. edodes. Lcc 1 was effective in the decolorization of chemically different dyes - Remazole Brilliant Blue R, Bromophenol Blue, methyl red, and Naphtol Blue Black - without any mediators, but the decolorization of two dyes - red poly(vinylamine)sulfonate-anthrapyridone dye and Reactive Orange 16 - did require some redox mediators.
Nagai
M
M
Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate 024-0003, Japan. nagai@ibrc.or.jp
Sato
T
T
Watanabe
H
H
Saito
K
K
Kawata
M
M
Enei
H
H
D003116:Color; D004396:Coloring Agents / Q000378:metabolism*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D042845:Laccase; D020074:Lentinula / Q000201:enzymology*; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
D004396:Coloring Agents; D008670:Metals; D010088:Oxidoreductases; D042845:Laccase