Purification and some properties of hemolysin produced by Corynebacterium pyogenes.

Hemolysin was purified from the culture supernatant of Corynebacterium pyogenes by successive ultrafiltration with PM 10 membrane, column chromatography on carboxymethyl cellulose and Sephadex G-200. The purified hemolysin was a substance of high molecular weight. This hemolysin formed only one band in polyacrylamide gel electrophoresis, but it gave about five bands in SDS-polyacrylamide gel electrophoresis. The hemolytic activity was not enhanced by the addition of L-cysteine, sodium hydrosulfite or sodium thioglycolate. Most of heavy metals did not inhibit nor stimulate the hemolytic activity. The hemolytic activity was completely destroyed by pronase or trypsin, but not by pepsin. It was inactivated by heating at 60°C for 10 minutes. Rabbit, horse and pig erythrocytes were most sensitive to the hemolysin and chicken erythrocytes were insensitive. The LD50 of hemolysin was approximately 96 hemolytic units per mouse.