The interaction of washed cocci, prepared under specified conditions, and a polynucleotide (AF) results in the formation of streptolysin S provided a fermentable carbohydrate is present. Maximum toxin formation requires, in addition, the presence of magnesium, potassium, and phosphate ions. Streptolysin S production proceeds anaerobically as well as aerobically but under the latter condition, apparently only if the system is sufficiently reducing. Temperature has a marked effect on the rate of appearance of toxin, the critical thermal increment having a value of approximately 36,000. The formation of streptolysin S is inhibited by mercuric ion, arsenite, iodoacetate, dinitrophenol, azide, and other enzyme poisons. The development of streptolysin S in resting cell systems depends neither upon autolysis nor upon physical extraction of preformed toxin but upon toxin synthesis.

From the supernatant fluid of the resting cell system, a product containing 20,000 to 30,000 units of streptolysin S per mg. dry weight can be isolated. Information concerning the pH stability of the product is presented. The product is free of streptokinase, hyaluronidase, and proteinase, but possesses appreciable desoxyribonuclease activity. Chemical analyses and other findings indicate that polynucleotide and carbohydrate are present in major amount, and that a small but undetermined quantity of protein is present. Inactivation of streptolysin S by chymotrypsin, ficin, papain, or cathepsin, and not by a variety of other enzymes, indicates that protein is essential for activity, but the precise chemical composition of the toxin remains to be established.

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