Purification and Some Properties of Cathepsin A of Small Molecular Size from Pig Kidney

Abstract

Cathepsin A [EC 3.4.2.-] of small molecular size (cathepsin A, S) has been purified about 800-fold from pig kidney by procedures including chromatographies on DEAE-Sephadex, SP-Sephadex, and Sephadex G-150.

1. The homogeneity of the purified enzyme was proved by ultracentrifugation and polyacrylamide gel elecrophoresis. The molecular weight (100,000) and isoelectric point (pI=5.0) were estimated.

2. The enzyme was remarkably stabilized by sucrose and KCl, and was most stable at pH 5–5.5 in the presence of both stabilizers. The enzyme had not only peptidase activity but also esterase and amidase activity; it was optimally active at pH 5.2 for peptide hydrolysis and at pH 8 for the hydrolysis of esters and amides.

3. Diisopropyl fluorophosphate and iodoacetamide completely inhibited these three activities.

4. The enzyme hydrolyzed various benzoyl- and benzyloxycarbonyl-dipeptides with neutral, acidic, and basic amino acids, and proline in the C-terminal position. The carboxypeptidase nature of the enzyme was proved by its action on an oligopeptide.

5. Several enzymatic properties of cathepsin A, S were almost the same as those of cathepsin A of large molecular size (cathepsin A, L) and the crude homogenate.