PROPERTIES AND ENZYMATIC ACTIVITIES OF PAPAIN INSOLUBILIZED WITH GLUTARALDEHYDE

Glutaraldehyde at a concentration of 2.3% and at 0 ° reacted in aqueous solutions with papain to form a water-insoluble product with enzymatic activity after activation by reducing agents. A rapid reaction of glutaraldehyde with the essential sulfhydryl of papain was not involved in the reaction since after activation the insoluble enzyme retained esterolytic and proteolytic activity. The amount of enzymatic activity retained by the insoluble material depended on the conditions of the reaction. The insolubilizatiou of papain with glutaraldehyde was pH dependent; the higher the pH the more rapid the reaction. Complete precipitation occurred in 24 hr over the range pH 5.2-7.2. The lower the pH of the reaction, the greater was the retention of esterase activities. Similar results were obtained when the reaction was carried out in the presence of added cysteine. However, the insoluble enzyme prepared in the absence of cysteine was fibrous while that prepared in its presence was crystalline. The reaction of mercuripapain with glutaraldehyde resulted in an insoluble enzyme that possessed, after treatment with cysteine, approximately twice as much esterase and proteinase activities as did the insoluble enzyme prepared from papain. Compared with soluble papain, preparations from glutaraldehyde mercuri-papain retained 48% of the ester, 12% of the casein, and 16% of the hemoglobin hydrolyzing ability. The reaction of the reduced soluble and insoluble enzymes with iodoacetamide-1-14C demonstrated that insoluble mercuripapain retained all of its essential sulfhydryl groups whereas those in insoluble papain had been reduced to two thirds. Amino acid analysis on hydrolyzed insoluble enzyme showed that only the lysine had been significantly altered by the glutaraldehyde reaction, having been reduced to approximately one-half that of soluble papain under all the conditions studied. © 1969.