A new affinity gel for purifing polyphenol oxidase enzyme

Abstract

Polyphenol oxidase (PPO) enzyme, sometimes called as phenoloxidase, catecholase, phenolase, catechol oxidase or tyrosinase, isconsidered to be an o-dipenol. PPO (EC 1.14.18.1), a multifunc-tional copper containing metalloenzyme, is widely distributed innature. PPO exists in many kinds of plants and fungi, such asbanana, mushroom, butter lettuce, Napoleon grape, potato, cof-fee, marula fruit, artichoke heads, longan fruit, tobacco, wheatflour.In this study, a novel affinity chromatography gel was synthe-sized for purifing PPO enzyme. The affinity chromatography gelwas synthesized by coupling aniline as a specer arm to CNBractiveted Sepharose-4B. Then, p-amino benzoic acid was coupledto aniline as a ligand. PPO was purified fromMusa sapientum var. Cavendishii(banana) by using Sepharose-4B-aniline p-aminobenzoic acid affinity chromatography gel. % 4.49 yield and 33.4fold purification were achieved. SDS-polyacrylamide gel elec-trophoresis of the enzyme indicates a single band with an appar-ent MW of 35 kDa. The Vmaxand Kmof the purified enzymewere determined 42,628 U/ml min and 9.27 mM, respectively.