Effect of Sulfhydryl and Other Reducin Compounds on De-carboxylation of Pyruvic Acid by Alkaline Washed Yeast. ∗

Since Grassman 1 first showed that papain and kathepsins can be activated by cysteine or by reduced glutathione many papers have appeared in the literature dealing with the influence of sulfhydryl compounds on the activity of hydrolyzing and other enzyme systems. A search of literature has not revealed any researches concerned with the effect of these compounds on carboxylase. This paper is a preliminary report on the effect of sulfhydryl and other reducing compounds on the activity of carboxylase.

The procedure used was a modification of the method of Ochoa and Peters. 2 As a source of carboxylase commercial baker's yeast which had been air-dried with the aid of an electric fan for 3-4 hours or desiccated over concentrated H₂SO₄ for several days was employed. It was freed from cocarboxylase by rapid washing of the yeast 2-3 times with M/10 Na₂HPO₄ (25-30 cc for 0.5 g yeast), and once with distilled water. The washed yeast was suspended in 5 cc M/10 phosphate buffer pH 6.2. Formation of CO₂ from pyruvic acid was measured manometrically at 28° in air with conventional Warburg manometers attached to 15 cc vessels. Each vessel contained 0.5 cc washed yeast suspension, 0.1 mg Mg as MgSO₄ in 0.1 cc of solution, 0.2 cc sodium pyruvate solution adjusted to pH 6.2 and containing 5 mg pyruvic acid. In each case reagents to be tested or phosphate buffer pH 6.2 were added to bring the total volume of fluid to 2 cc. The pyruvate solution was tipped into the vessel after 8-12 minutes aeration. Merck's synthetic cocarboxylase and crystalline vitamin B₁, were used throughout the experiments.†

The results of 5 experiments dealing with the activation of alkaline washed yeast by cysteine hydrochloride, reduced glutathione, sodium bisulfite and phenylhydrazine hydrochloride are listed in Table I.