Egg Capsule Catechol Oxidase from the Little SkateRaja erinaceaMitchill, 1825

A phenoloxidase was demonstrated in extracts of egg capsules tanning in utero and of nidamental glands from spawning little skate, Raja erinacea. The enzyme was identified as a catechol oxidase based on its ability to oxidize the ortho-diphenols pyrocatechol, 4-methylcatechol, 3,4-dihydroxyphenylalanine, 3-hydroxytyramine and N-acetyldopamine to their corresponding ortho-quinones and its relative inactivity against monophenols. 4-methylcatechol was oxidized at the greatest rate, while 3,4-dihydroxyphenylalanine, 3-hydroxytyramine and N-acetyldopamine were oxidized at slower rates. The nidamental gland enzyme was inhibited by cyanide, nitrogen, and diethyldithiocarbamate. Oxidase activity in crude extracts from nidamental glands was enhanced by addition of α-chymotrypsin, suggesting that the enzyme is produced in a latent form. Ammonium sulfate fractionation of nidamental gland and capsule extracts resulted in a fifteen-fold purification of the enzyme. This partially purified catechol oxidase from the nidamental gland exhibited optimal rates of oxidation at 0.5 M NaCl and pH 7.0. The enzyme, however, showed a wide tolerance for elevated salinity and alkaline pH. These observations indicate that the oxidase acts principally in utero, but may remain active in seawater following oviposition of the capsule. This enzyme plays a pivotal role during the formation of skate egg capsules by catalyzing the oxidation of capsular catechols to highly reactive quinones forming dark pigments which tan the capsular matrix.