Improved purification and further characterization of acid carboxypeptidase from Aspergillus saitoi.

Abstract

An acid carboxypeptidase was highly purified and characterized from Aspergillus saitoi ATCC 14332. By disc gel electrophoresis at pH 9.4, SDS disc gel electrophoresis, and isoelectric focusing, the enzyme was essentially homogeneous. The molecular weight was 125,000 by gel filtration and 72,000 by SDS disc gel electrophoresis. The isoelectric point was 4.07. The enzyme acted as a carboxyamidase for cholecystokinin-tetrapeptide (CCK-tetrapeptide, Trp-Met-Asp-Phe-NH2). The Km and Kcat values of the enzyme for Z-Glu-Tyr, angiotensin, and bradykinin at pH 3.1 and 30°C were 4.0mm and 106 sec-1, 0.05mm and 0.35 sec-1, and 0.04mm and 6.0 sec-1, respectively. An aqueous solution of the enzyme was freeze-dried retaining 98% of the enzyme activity. None of the activity was lost after preservation for 3 years at 4°C.