PROPERTIES OF PARTIALLY PURIFIED PIG BRAIN STEM TRYPTOPHAN HYDROXYLASE

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Abstract
—Tryptophan hydroxylase form pig brain has been purified using a method which involved sonic disintegration of a whole homogenate, ammonium sulphate fractionation, hydroxylapatite fractionation, column chromatography on Sephadex G‐100 or G‐200 and finally electrophoresis on poly‐acrylamide gel. The enzyme was stabilized during purification by tryptophan and dithiothreitol. The partially purified enzyme has a molecular weight of 55,000‐60,000 as measured by gel‐filtration. The Km of the soluble partially purified enzyme was 0‐4 mm, which differed significantly from that of the particulate enzyme (0·02mm). Enzyme activity was not stimulated by ferrous ion. However, it was inhibited by the chelating agents 8‐hydroxyquinoline, O‐phenanthroline and EDTA. In contrast to dopamine, high concentration of tryptophan (10 mm), 5‐hydroxytryptamine, tryptamine and tyramine at 0‐5 mm concentration did not inhibit the enzyme in the presence of dimethyltetrahydropterin (DMPH4). A number of monoamine oxidase inhibitors, phenelzine, pheniprazine and chlorgyline at 1 mm strongly inhibit the formation of 5‐hydroxytryptamine. Evidence is presented for the presence of an endogenous inhibitor of tryptophan hydroxylase.

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