Purification and partial characterization of the pancreatic proteolytic enzymes trypsin, chymotrypsin, and elastase from the chicken.
The purpose of this work was to isolate, purify and partially sequence trypsin, chymotrypsin and elastase from the chicken pancreas. The extraction of the pancreatic zymogens with 0.5 M CaCl2 at pH 7.5 for 9 h appeared to be most effective in obtaining maximum recovery of the three enzymes. The sequential Cucurbita maxima trypsin inhibitor I/bovine pancreas trypsin inhibitor/soybean trypsin inhibitor affinity chromatography gave the best result for the isolation of trypsin, chymotrypsin and elastase, respectively, from the same extract. For each proteinase, multiple form of enzymatic activity could be observed after gel electrophoresis and each form was further purified on an ion-exchange column. The N-terminal amino acid sequence of trypsin and chymotrypsin showed homologies with the bovine enzymes whereas elastase showed homologies with the porcine enzyme. The molecular mass of trypsin, chymotrypsin and elastase were estimated to be 23,500, 25,700 and 25,000, respectively, which are values close to those in mammalian species. Although some kinetic constants (Km and k(cat)/Km) appeared different from those observed in other species, the pH dependent enzymatic activities were similar to those reported in other animal species.
Guyonnet
V
V
Department of Poultry Science, University of Georgia, Athens 30602, USA.
Tłuscik
F
F
Long
P L
PL
Polanowski
A
A
Travis
J
J
D000818:Animals; D002645:Chickens; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D002918:Chymotrypsin / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology*; D010196:Pancreatic Elastase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014357:Trypsin / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
D002918:Chymotrypsin; D010196:Pancreatic Elastase; D014357:Trypsin