Large Scale Production and Crystallization of Acid Carboxypeptidase from Submerged Culture of Penicillium janthinellum, and Stability of the Crystalline Enzyme

Abstract

Acid carboxypeptidase of Penicillium janthinellum IFO-8070 was produced effectively in submerged culture on a medium of 4_??_5% rice bran. The enzyme production was enlarged to volume cultivation of 150-liters in a 200-liters jar fermentor, and the yield of acid carboxy-peptidase per milliliter filtrate reached to the maximum 3 days after inoculation.
Acid carboxypeptidase of low molecular weight (M. W.=51, 000) produced in the liquid culture broth was purified and crystallized in a large scale. Purification steps include Amberlite CG-50 treatment, ammonium sulfate precipitation, dialysis using “Diaflow, ” activated charcoal treatment, and condensation using collodion-bag, or condensation and dialysis using “Diaflow.”
The crystals of the acid carboxypeptidase suspended in 50mM acetate buffer (pH 3.7) were completely stable for six months at 5°C. On the other hand, at low enzyme concentration (0.01 U/ml) in 50mM acetate buffer (pH 3.7), crystallized enzyme was somewhat labile, whereas, this inactivation was completely depressed by covering enzyme solution with toluene.