Purification and characterization of the pancreatic proteolytic enzymes trypsin and chymotrypsin from the Kamori goat of Sindh, Pakistan, using ion-exchange chromatography

• Published in: Separation science and technology Vol. 60; no. 3-5; pp. 467 - 479
• Main Authors: Khan, Ariba, Talpur, Farah Naz, Bhanger, Muhammad Iqbal, Musharraf, Syed Ghulam, Afridi, Hassan Imran, Qambrani, Shagufta
• Format: Journal Article
• Language: English
• Published: Abingdon Taylor & Francis 24.03.2025; Taylor & Francis Ltd
• Subjects: Arginine; Calcium; Chromatography; Chymotrypsin; Enzymes; Ethyl esters; Goats; Ion exchange; Ion-exchange chromatography; Kamori goat; Molecular weight; Pancreas; pH effects; Proteolysis; Proteolytic enzymes; Slaughterhouse waste; Sodium chloride; Stability; Substrate inhibition; Trypsin; Tyrosine; Water purification; Sindh Pakistan; Pakistan
• ISSN: 0149-6395; 1520-5754
• DOI: 10.1080/01496395.2024.2436480

Trypsin and chymotrypsin were purified from the Kamori goat pancreas by affinity and ion exchange chromatography, and their activity was checked. The activity and stability of extracted enzymes under various conditions, including temperature, pH, and substrates, were studied. Extracted enzyme's activity and stability studies demonstrated that both enzymes possessed excellent activity at pH 8 and reached maximum activity after 8 hours. Purity was increased to 22.1-and 12.8-fold, with 29.2% and 25.3% yields for trypsin and chymotrypsin, respectively. According to SDS-PAGE, their molecular weights were approximately 24 and 25.5 kDa, respectively. Trypsin and chymotrypsin displayed maximum activity at 50°C, pH 8, using BAPNA (Na-benzoyl-DL-arginine 4-nitroanilide hydrochloride) and BTEE (N benzoyl-L-tyrosine ethyl ester) as substrates. Both enzymes were stabilized in the presence of Ca ++ ions. The enzymes activity declined continuously as the concentration of NaCl increased. Enzymes were effectively inhibited by SBTI and PMSF but not by pepstatin A. K cat of trypsin and chymotrypsin was 16.8 and 36.7 S −1 , and K m was 0.91 and 0.17 mm, respectively. Results suggest that Kamori goat pancreas is a promising source of trypsin and chymotrypsin due to their excellent activity and stability.