Purification of Yeast Proteinases

Abstract

Three types of proteinases contained in baker’s yeast were fractionated and partially purified by chromatography on TEAE-cellulose and on DEAE-Sephadex. These enzymes were designated as proteinase A, Band C, and some properties of proteinase A and B were described. A was similar to pepsin and “acid-proteinase” produced by various fungi, but did not hydrolyze carbobenzoxY-L-glutamyl-L-tyrosine. B had milk-clotting activity and proteolytic activity toward casein, and also esterolytic activity toward N-acetyl-L-tyrosine ethylester and α-N-benzoyl-L-arginine ethylester. This enzyme was inhibited by p-mercuribenzoate and diisopropylphosphorofluoridate. Yeast proteinase C which shows strong esterolytic activity against N-acetyl-L-tyrosine ethylester was successfully purified from baker’s yeast. After repeated chromatography on TEAE-cellulose column, a single elution pattern was obtained regarding protein and the esterolytic activity. The proteolytic activity could not be measured in 0.5% casein solution, but was observed clearly by the use of 4% casein solution as the substrate. Both of the proteolytic activity and the esterolytic activity were inhibited by the sulfhydryl reagents such as p-mercuribenzoate and also by diisopropylphosphorofluoridate.