title: Purification and characterization of two pepsins from the stomach of pectoral rattail (Coryphaenoides pectoralis).
abstract: Two pepsins (A and B) were purified from the stomach of pectoral rattail (Coryphaenoides pectoralis) by acidification, ammonium sulfate precipitation, gel filtration chromatography and anion exchange chromatography to obtain a single band on native-PAGE and SDS-PAGE. The purities of pepsin A and B were increased to 7.1- and 13.0-fold with approximately 5.7% and 2.2% yield, respectively. Pepsin A and B had the apparent molecular weights of 35 and 31 kDa, respectively, when analyzed using SDS-PAGE and Sephacryl S-200 gel filtration. Pepsin A and B showed maximal activity at pH 3.0 and 3.5, respectively, and had the same optimal temperature at 45 degrees C using hemoglobin as a substrate. Both pepsin A and B were stable in the pH range of 2.0-6.0 but were unstable at the temperatures greater than 40 degrees C. Activity of both pepsins was inhibited by pepstatin A and was activated by divalent cations, indicating pepsin characteristics. Activities of both pepsins continuously decreased as NaCl concentration increased (0-30%). The enzymes had high affinity and activity toward hemoglobin with Km and Kcat values of 98-152 microM and 32-50 S(-1), respectively. Purified pepsins generally showed the similar characteristics to other fish pepsins.
authors.affiliation: Department of Food Science and Technology, Faculty of Technology and Community Development, Thaksin University, Phattalung Campus, Phattalung, 93110, Thailand. sappasith@tsu.ac.th
authors.forename: Sappasith
authors.initials: S
authors.lastname: Klomklao
authors.forename: Hideki
authors.initials: H
authors.lastname: Kishimura
authors.forename: Mamoru
authors.initials: M
authors.lastname: Yabe
authors.forename: Soottawat
authors.initials: S
authors.lastname: Benjakul
mesh_terms: D000818:Animals; D048388:Gadiformes; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D010434:Pepsin A / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D011489:Protein Denaturation / Q000187:drug effects; D012965:Sodium Chloride / Q000494:pharmacology; D013270:Stomach / Q000737:chemistry*; D013696:Temperature
chemical_list: D012965:Sodium Chloride; D010434:Pepsin A; C095924:pepsin B