Thermophilic laccase from xerophyte species Opuntia vulgaris.
Two laccase temperature isoforms capable of oxidizing phenolic compounds to quinones were isolated and purified to homogeneity from the cladodes of the xerophyte species Opuntia vulgaris. These catalytically active proteins exhibit apparent molecular masses of 137 and 90 kDa. Under reducing conditions, both isoforms yielded a subunit molecular mass of 43 kDa, suggesting that the enzyme is a multimer of the 43 kDa subunit. The 137 kDa isoform when heated at 80°C for 3 min generated three polypeptide bands on activity stained polyacrylamide gels exhibiting 137, 90 and 43 kDa molecular forms. All isoforms of the enzyme exhibited an optimum pH of 10 when 2,6-dimethoxyphenol was used as a substrate. The optimum temperature of the 137 kDa enzyme form was noted to be 80°C and that of the 90 kDa enzyme form was 70°C. Denaturation kinetics of both the laccase isoforms carried out at their respective optimum temperatures for 30 min exhibited enzyme activity in excess of their t(1/2) values throughout the assay period. The K(m) for the 137 kDa form was determined to be 2.2 ± 0.3 mm and the V(max) was 2.8 ± 0.2 IU/mL. These high temperature stable laccase isoforms having alkaline pH optima can find significant industrial use.
Kumar
Gali Nirmal
GN
Department of Biochemistry and Molecular Biology, School of Life Sciences, Pondicherry University, Puducherry, India.
Srikumar
Kotteazeth
K
D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D042845:Laccase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D029747:Opuntia / Q000201:enzymology*; D010936:Plant Extracts / Q000737:chemistry; D018547:Plant Stems / Q000737:chemistry; D011489:Protein Denaturation; D021122:Protein Subunits; D011748:Pyrogallol / Q000031:analogs & derivatives / Q000737:chemistry
D010936:Plant Extracts; D021122:Protein Subunits; D011748:Pyrogallol; C010120:pyrogallol 1,3-dimethyl ether; D042845:Laccase