Serum phenoloxidase activity in the hemolymph of the anomuran crab Albunea symmysta (Linnaeus, 1758) (Decapoda: Anomura: Albuneidae)

Abstract

Abstract We characterized the optimal conditions for measuring serum phenoloxidase activity and its functional activity and susceptibility to an inhibitor and various activators in an anomuran crab, Albunea symmysta (Linnaeus, 1758). The substrate affinity of the phenoloxidase (PO) enzyme was determined using different phenolic substrates in which only diphenols were found to be oxidized. The enzyme was characterized as a catecholoxidase-type of PO and 3,4-dihydroxy-DL-phenylalanine (DL-Dopa), the enzyme showing the highest substrate affinity to the serum. The optimal enzyme activity was observed at 5 mM DL-Dopa in 10 mM Tris-HCl buffer at a pH of 7.5 at 25 °C for 10 min, and absorbance at 470 nm. Serum-PO activity was inhibited by 7 mM phenylthiourea (PTU), and activated by activators such as trypsin, chymotrypsin, pronase-E, and detergent-like sodium dodecyl sulfate (SDS). We also identified the chemicals causing in vitro inhibition or activation of the enzyme as a serum of the crab having a potent PO activity.