International Research Journal of Commerce , Arts and Science

 ( Online- ISSN 2319 - 9202 )     New DOI : 10.32804/CASIRJ

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PARTIAL PURIFICATION AND CHARACTERIZATION OF LACCASE FROM PLEUROTUS OSTREATUS

    6 Author(s):  CHANDRASHEKAR S,SOWMYA H.K., CHANDAN S, SHIVA PRASAD K.,G. PANDURANGA MURTHY,SHIVALINGAIAH

Vol -  11, Issue- 1 ,         Page(s) : 46 - 69  (2020 ) DOI : https://doi.org/10.32804/CASIRJ

Abstract

Laccases are either mono or multimeric copper-containing oxidases that catalyze the one electron oxidation of a vast number of phenolic substrates. Molecular oxygen serves as the terminal electron acceptor and is thus reduced to two molecules of water. The ability of laccases to oxidize phenolic compounds and reduce molecular oxygen to water has led to intensive study of these enzymes. Laccase is of interest in biotechnology because of the reactions that it is able to catalyze. A number of industrial applications for laccases have been proposed and they include paper processing, prevention of wine decolouration, detoxification of environmental pollutants, oxidation of dye and dye precursors, enzymatic conversion of chemical intermediates and the production of valuable compounds from lignin. The ideal laccases for industrial use would exhibit stability at high temperature and pH conditions. Laccase enzyme was identified and isolated from Pleurotus sp. The final purification protocol consisted of precipitation with ammonium sulphate at 80% saturation and dialysis. The final purification led to a homogenous product with a specific activity of 848 U/mg and a purification yield of 86%. Laccase from Pleurotus sp. has a molecular mass of approximately 55±1 KDa. The partially purified protein exhibits a temperature optimum of 65°C. The optimum pH differed for different substrates and was evaluated for ABTS (pH 4.5) and Syringaldazine (pH 6.0). Kinetic analysis for Plerotus species included evaluation of two substrates. The Km values for ABTS and 2, 6-DMP were 250mM and 38.46mM respectively and the Vmax values were 0.33μmol/min and 20μmol/min respectively. Inhibition studies for the partially purified enzyme included two different inhibitors (sodium azide and EDTA). The Ki values were 0.03 mM for sodium azide and 3.2mM for EDTA. The purified laccase showed potent antimicrobial activity against E.coli at 14 µl.

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