Journal article
Purification and characterization of two chymotrypsin-like proteases from the pyloric caeca of rainbow trout oncorhynchus-mykiss
Two chymotrypsins, called chymotrypsin I and II, were purified from the pyloric caeca of rainbow trout, by (NH4)2SO4 fractionation, hydrophobic interaction chromatography (phenyl-Sepharose) and ion-exchange chromatography (DEAE-Sepharose). The approximate molecular weights of chymotrypsin I and II were 28,200 (.+-.1200) and 28,800 (.+-.900), respectively, as determined by SDS-PAGE and their isoelectric points were about 5.
The pH optima of the enzymes were centered around nine, when assayed for succinyl-L-Ala-L-Ala-L-Pro-L-Phe-p-nitroanilide (Such-AAPF-NA) as substrate and both enzymes were unstable at pH values below 5. The amidase activity of both enzymes increased with temperature up to about 55.degree. C. Chymotrypsin I was found to be more heat stable than chymotrypsin II, an effect most likely explained by strong calcium binding of the former.
The trout chymotrypsins were significantly more active than bovine .alpha.-chymotrypsin when assayed against Suc-AAPF-NA at 25.degree. C and casein at low temperatures (10-20.degree. C), indicating an adaptation of the activities of the trout chymotrypsins to the habitation temperatures of the fish.
Language: | English |
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Year: | 1992 |
Pages: | 247-253 |
ISSN: | 18791107 and 10964959 |
Types: | Journal article |
07508, Ecology: environmental biology - Animal 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines 10802, Enzymes - General and comparative studies: coenzymes 10804, Enzymes - Methods 10806, Enzymes - Chemical and physical 14004, Digestive system - Physiology and biochemistry 23012, Temperature - Thermoregulation 9014-01-1D: PROTEASES7783-20-2: DIAMMONIUM SULFATE Amino Acid Sequence Animals Biochemistry and Molecular Biophysics Calcium Caseins Catalysis Cattle Chromatography, Gel Chymotrypsin DIAMMONIUM SULFATE FRACTIONATION ION-EXCHANGE CHROMATOGRAPHY HYDROPHOBIC INTERACTION CHROMATOGRAPHY THERMOREGULATION Digestive System EC 3.4.21.- Serine Endopeptidases EC 3.4.21.1 Chymotrypsin Ecology Electrophoresis, Polyacrylamide Gel Environmental Sciences Enzyme Stability Enzymology Hydrogen-Ion Concentration Ingestion and Assimilation Intestines Molecular Sequence Data Physiology Pisces Vertebrata Chordata Animalia (Animals, Chordates, Fish, Nonhuman Vertebrates, Vertebrates) - Osteichthyes [85206] SY7Q814VUP Calcium Salmon Serine Endopeptidases Serine Proteinase Inhibitors Temperature