Isolation and characterization of the polyphenoloxidase from senescent leaves of black poplar |
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Authors: | Michèle Trémolières Joseph G Bieth |
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Affiliation: | 1. Laboratoire de Botanique et Laboratoire d''Enzymologie, INSERM Unite 237 Strasbourg Cedex, France;2. Faculté de Pharmacie, Université Louis Pasteur, BP 10 67048 Strasbourg Cedex, France |
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Abstract: | The polyphenoloxidase (PPO) from black poplar senescent leaves has been purified to almost complete homogeneity by a combination of ammonium sulphate precipitation, Sephadex G75 filtration and DEAE-cellulose chromatography. The purified enzyme has a MW of 60 000 and is probably a Cu+ enzyme. Peroxidase (PO) activity co-purifies with PPO and has the same MW as it. The two enzymes differ in pH optimum and in response to the effect of ionic strength. Natural phenols are either substrates, inhibitors or activators of black poplar PPO. This enzyme is an o-diphenoloxidase which binds substrates with Km in the millimolar range. With caffeic and chlorogenic acids inhibition by excess substrate is observed. Benzoic acid phenols and cinnamic acid phenols are either competitive or non-competitive inhibitors of PPO. Hydroquinone is a highly potent non-competitive inhibitor of the enzyme (Ki 90 μM). Ferulic acid is a potent activator of the PPO-catalysed oxidation of catechol (Ka 0.34 mM, νsat/νo 7.7). |
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Keywords: | black poplar Salicaceae polyphenoloxidase peroxidase senescent leaves transition metals oligoelements phenols inhibition activation |
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