Methods.-(1) Enzyme essays: One unit of each of the enzymatic activities followed is the amount of enzyme catalyzing formation of 0.1 Mole of product in 10 min at 37. The specific activity is expressed as the number of units of enzyme per milligram of protein. Solutions of substrate amino acids were adjusted to pH 7.8 with KOH before addition to assay solutions. Assays for individual enzymatic activities were conducted as follows:(a) TPase: The procedure described previously'was modified slightly by increasing the potassium phosphate buffer (pH 7.8) from 20to 40 Mmoles, and adding 0.25 mg of bovine serum albumin.(b) TSase: The assay described previously1 was used with the following changes: thebuffer consisted of 20, umoles of potassium phosphate, pH 7.8, the reaction was started by addition of 32 lumoles of L-serine, and was allowed to proceed for 10 min. The disappearance of indole and the formation of tryptophan were followed by use of the acid-Ehrlich reagent and by microbiological assay with Leuconostoc mesenteroides P-60, respectively. Synthesis of tryptophan from L-cysteine and S-methyl-icysteine was followed by microbiological assay of reaction mixtures containing the appropriate amino acid as substrate in place of iserine.(c) DeaminationofLnserine, L-cysteine, and S-methyl-cysteine: These reactionswere followed by measuring pyruvate formation in 0.2 ml of reaction mixture containing 20 pmoles of potassium phosphate, pH 7.8, 20 miumoles of pyridoxal phosphate, 32 jsmoles of the amino acid, and enzyme. After 5 min at 370, the reaction is started by addition of the amino acid and allowed to proceed for 10 min with shaking. The reaction is stopped by addition of 0.1 ml of 2.5 N sodium hydroxide; then 0.5 ml of 2, 4-dinitrophenylhydrazine in 2 N hydrochloric acid is added. After 5 min, 1.5 ml of absolute ethanol is added and the c'Aor developed by the addition of 2.5 ml of 2.5 N sodium hydroxide. After 10 min, the absorbance at 520 mu is determined, and the amount of pyruvate formed is calculated froma standard curve prepared using crystalline sodium pyruvate. The sulfhydryl group interferes with estimation of pyruvate when cysteine is the substrate. This was prevented by addition of 64 jmoles of iodoacetamide in 0.2 ml of water to the reaction mixture immediately after stopping the reaction with alkali and by allowing the mixture to stand for 5 min before adding the 2, 4-dinitrophenylhydrazine reagent.(2) Other analytical procedures: Hydrogen sulfide in reaction mixtures was trapped as cadmium sulfide in the center well of a Warburg vessel and determined as described by Smythe.'The incubation was carried out for 20 min at 370 with shaking. Ammonia was determined after the reaction was stopped with 2 N hydrochloric acid (0.5 ml addedto a reaction volume of 2 ml) by adsorption and elution from permutite followed by Nessler's reagent. 4 Starch gel electrophoresis patterns were obtained by a modification of the procedure of Barrett, Friesen, and Astwood. 5 The TPase preparations were diluted to 10 mg protein per ml and dialyzed overnight against 0.01 M potassium phosphate buffer (pH 7) before applying to the gel.(3) Culturesand growth media: Mutantstrains B/1t7 and B/1t7-A of E. coli and media for their culture have been described. 1 Mutant B/1t7-A, which produces tryptophanase constitutively, was grown at 300 with vigorous aerationin 16-1 carboys. Each carboy, containing 121 of broth medium, was inoculated with 41 of an overnight culture grown on minimal medium supplemented with 0.1% glucose and10 p&g/ml of indole. The cells were harvested by centrif-ugation at the end of the logarithmic phase of growth (4-6 hr …