Tyrosinase In Crustacean Blood

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Abstract
The blackening of crustacean blood when it is shed, or in clots at wounds, is caused by an enzyme similar to, if not identical with, the tyrosinase systems previously described in various invertebrates, fungi, and in the higher plants. The components of the system are an enzyme contained in the blood corpuscles, from which it is freed on cytolysis, and its substrate tyrosine, which is free in the blood stream. The enzyme is by definition a tyrosinase, since it will bring about the oxidation of tyrosine with the ultimate production of melanin, deriving the oxygen necessary for the reaction from the air. The tyrosinase content of the blood is not constant, nor does it undergo a seasonal variation. The possibility of shorter cycles in tyrosinase activity has not been investigated. The blood will accelerate the oxidation of diphenylenediamine and α napthol to the blue indophenol derivative; but as this reaction is comparatively insensitive to NaCN, it is unlikely that it is due to an indophenol oxidase1 and is probably caused by the autoxidation of some substance in the blood such as an orthoquinone. The tyrosinase action is inhibited by low molecular concentrations of NaCN, indicating the presence of a metallic group as the active part of the enzyme molecule. The activity of the enzyme can be depressed by H2S, CuSO4 FeCl3 sodium fluoride, sodium pyrophosphate, and the alcohols. Of the latter, ethyl alcohol is effective in concentrations of 2.7 molar, while methyl alcohol tested on the same blood had no effect in concentrations of 3.5 molar. This is the expected result from Warburg's hypothesis, but as tyrosine is insoluble in alcohols, and the amounts of the higher alcohols which could be introduced into the watery solution were too small to have any effect on the oxidation, a series could not be investigated. Thymol, phenyl urethane and urethane will depress the oxidation of tyrosine by the enzyme. The anomalous effect of certain concentrations of these reagents in increasing the rate and amount of oxidation is discussed.