Biochemical properties of polyphenoloxidase from the cephalothorax of Pacific white shrimp (Litopenaeus vannamei)

Partially purified PPO (83.8-fold purity) showed the maximal activity using L-β-(3,4-dihydroxylphenyl)alanine (L-DOPA) as a substrate at pH 6 and at 55°C. PPO was stable over a pH range of 5 to 10 but unstable at a temperature greater than 60°C. Based on the activity staining with L-DOPA, the apparent molecular weight of PPO was 210 kDa. The Michaelis constant (Km) of PPO for the oxidation of L-DOPA was 2.43 mM. Trypsin, copper acetate and sodium dodecyl sulphate were unable to activate PPO, suggesting that the enzyme was in the active form. Cysteine, ethylenediaminetetraacetic acid and p-aminobenzoic acid showed PPO inhibitory activity in a dose-dependent manner. At the same concentration used (1 and 10 mM), cysteine exhibited a higher inhibitory effect towards PPO.Active PPO with higher stability could be responsible for the rapid melanosis formation in Pacific white shrimp, especially at the cephalothorax portion during postmortem storage. The use of an effective PPO inhibitor, especially cysteine, in combination with chilled storage could prevent melanosis in shrimp.Polyphenoloxidase (PPO) is known as phenolase, tyrosinase and catechol oxidase. PPO is a copper-containing metalloenzyme, which catalyses two basic reactions, in the presence of molecular oxygen, that include the o-hydroxylation of monophenols to give o-diphenols (monophenol oxidase, cresolase activity, EC 1.14.18.1) and the subsequent oxidation of o-diphenols to o-quinones (diphenoloxidase, catecholase activity, EC 1.10.3.1) (Garcia-Molina et al. [2005]). PPO in crustaceans is distributed mainly in the carapace, followed by the abdominal exoskeleton, cephalothorax, pleopods and telson (Zamorano et al. [2009]). PPO is most commonly found in the cephalothorax of prawn and shrimp (Montero et al. [2001a]). PPO is involved in the black spot formation in crustaceans during postmortem storage. Black spot formation (melanosis) is one of the problems that occur in crustaceans. It drastically reduces the cons