Purification of carboxypeptidase A using sepharose 4B-bound 3-phenylpropionate

• Published in: Journal of biochemistry (Tokyo) 1977-05, Vol.81 (5), p.1285-1291
• Main Authors: Oshima, G, Nagasawa, K. (Kitasato Univ., Tokyo (Japan). School of pharmaceutical Sciences)
• Format: Article
• Language: English
• Subjects: Carboxypeptidases - isolation & purification; Carboxypeptidases - metabolism; Chromatography, Affinity - methods; Kinetics; Pancreas - enzymology; Phenylpropionates; Structure-Activity Relationship
• ISSN: 0021-924X; 1756-2651; 1756-2651
• DOI: 10.1093/oxfordjournals.jbchem.a131581

The activity of carboxypeptidase A [EC 3.4.12.2] was inhibited by 3-phenylpropionate derivatives (p-aminocinnamate, 3-p-aminophenylpropionate and 3-p-acetylaminophenylpropionate), and to investigate its use as a ligand for affinity chromatography 3-p-aminophenylpropionate was directly and indirectly coupled to Sepharose 4B. Carboxypeptidase A was adsorbed only on 3-p-aminophenylpropionate bound to the gel through p-phenylenediamine as a spacer. Carboxypeptidase A from pancreas was purified by a combination of this affinity adsorbent and ion exchange chromatography. The purified carboxypeptidase A had a homogeneity similar to that of a commercial product, as judged by disc gel electrophoresis. The carboxypeptidase activity of Pronase was slightly retarded on the gel column, but could not be separated from its caseinolytic activity. Angiotensin I-converting enzyme [peptidyl dipeptide hydrolase, EC 3.4.15. 1] obtained from hog kidney cortex was not bound to the gel.