Protyrosinase and Polar-Nonpolar Cations and Anions. ∗

The sodium alkyl sulfates, which activate protyrosinase, 1 yield polar-nonpolar anions, (RSO₄). Alkylamine hydrochlorides, however, give polar-nonpolar cations (RNH₃.H⁺), and do not activate protyrosinase. † This distinction may be of value in the search for the kind of chemical group or part of protyrosinase which is involved in its activation.

Three similar preparations of protyrosinase were extracted from grasshopper eggs (Melanoplus differentialis) by a method involving trituration, fractional precipitation, and dialysis. 1 Activities, measured in a Warburg apparatus, were compared by the reciprocal—multiplied by 1000—of the time in minutes for the first 100 μ1 of oxygen uptake with tyramine hydrochloride as substrate.

The ascending limb of curve A (Fig. 1) shows the activation of protyrosinase that occurs in the presence of sodium dodecyl sulfate. With dodecylamine hydrochloride (Curve F, Fig. 1) there seems to be no conversion of protyrosinase into tyrosinase. When solutions of alkyl sulfates and alkylamines are mixed, a precipitate forms. Thus, from activation experiments performed by adding a constant amount of protyrosinase to such mixtures (Curves B, C, D, E, Fig. 1), it was found at half activation that 1.00 mole of either decylor dodecylamine hydrochloride removes 0.36, 1.01, and 2.79 moles of sodium octyl, decyl, and dodecyl sulfates (Fig. 2). These values may well be a function of the dissociation constants of the compounds formed by cations and anions.

The effects of variation in the order of mixing of reagents is graphically illustrated in Fig. 3. The upper curve (A) shows only a slight decrease in tyrosinase when excess dodecylamine is added after the mixing of protyrosinase and dodecyl sulfate. Curve B describes the effect, previously noted, of mixing dodecyl sulfate with dodecylamine and, after equilibration, adding the protyrosinase. Curve C refers to the results of mixing protyrosinase and dodecylamine and then adding the dodecyl sulfate.