ISOLATION, CHARACTERIZATION AND POTENTIAL APPLICATION OF DEOXYRIBONUCLEASE-FREE PHOSPHATASE FROM CASSAVA LEAVES

Autor:	Hock-Hin Yeoh, Saw Hoon Lim, S. C Tham
Rok vydání:	2006
Předmět:	chemistry.chemical_classification Chromatography biology Chemistry Chromatofocusing Phosphatase Acid phosphatase Horticulture Enzyme assay Enzyme Enzyme inhibitor biology.protein Alkaline phosphatase Ammonium sulfate precipitation
Zdroj:	Acta Horticulturae. :263-268
ISSN:	2406-6168 0567-7572
DOI:	10.17660/actahortic.2006.703.34
Popis:	A deoxyribonuclease-free acid phosphatase from cassava leaves was prepared by ammonium sulfate precipitation, chromatofocusing and hydrophobic interaction chromatography with phenyl Sepharose. The enzyme was purified 36-fold and had a specific activity of 16 U/ mg protein. The enzyme preparation revealed a major phosphatase band of 77 kDa and three minor activity bands. The pH and temperature optima for enzyme activity were 5.2 and 60°C, respectively. The enzyme was inactivated by heating at 80°C for 15 minutes and could be stored at -20°C for up to two months. The enzyme exhibited broad substrate specificity and had a K m (p-nitrophenyl phosphate) value of 1.7 mM. It was strongly inhibited by zinc and copper ions, molybdate and arsenate. Aluminium ions, ferrous ions, dithiothreitol and mercaptoethanol stimulated enzyme activity within 0.5-5.0 mM range. The enzyme partially dephosphorylated linearized pUC18 plasmid despite being an acid phosphatase and showed potential application as a substitute for alkaline phosphatase in the ELISA procedure.
Databáze:	OpenAIRE
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