Characterization of tyrosinase and accompanying laccase from Amorphophallus campanulatus.
Tyrosinase and laccase activities were detected in the corm of Amorphophallus campanulatus after extraction with ethanol followed by ammonium sulphate precipitation (20-60%) and dialysis against 10 mM Na2HPO4 buffer at pH 7.0. Tyrosinase was found to be the predominant enzyme exhibiting mono- and di-phenolase activities, specificity for L-DOPA as substrate, optimum pH being 6.0, optimum temperature at 40 degrees C and Km at 1.05 mM. Laccase showed substrate specificity for p-phenylenediamine (p-PD), Km at 2.7 mM, optimum pH being 5.0 and was inactivated above 40 degrees C. Three isoforms of tyrosinase were detected on SDS-PAGE with apparent molecular mass approximately 127, 31 and 27 kDa respectively. On staining sections of A. campanulatus with L-DOPA as substrate and 3-methyl benzothiazolinone hydrazone (MBTH) for colour development, tyrosinase was detected in the intercellular spaces of the plant tissue. The cytosolic region did not show any colour indicating the absence of the enzyme.
Department of Chemistry, University of Pune, Pune 411007, India.
Pallavi S
PS
Paranjpe
Meena S
MS
Karve
Subhash B
SB
Padhye
D031056:Amorphophallus / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D042845:Laccase / Q000378:metabolism*; D015416:Maillard Reaction; D014442:Monophenol Monooxygenase / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D021381:Protein Transport; D013379:Substrate Specificity; D013696:Temperature
D004791:Enzyme Inhibitors; D042845:Laccase; D014442:Monophenol Monooxygenase