Zymogen activation: effect of peptides sequentially related to the bovine beta-trypsin N-terminus on Kazal inhibitor and benzamidine binding to bovine trypsinogen.
The activating effect of peptides sequentially related to the Ile 16-Val17-Gly18 N-terminus of bovine beta-trypsin (namely Ile-Val-Gly, Ile-Val, Ile-Leu, Ile-Ala, Val-Val, Leu-Val, and Val-Leu) on the thermodynamic parameters for the binding of the porcine pancreatic secretory trypsin inhibitor (Kazal inhibitor) and benzamidine to bovine trypsinogen was investigated at pH 5.5 (Bis tris-HCl buffer, I = 0.1 M) and T = 21 +/- 0.5 degrees C. Thermodynamic parameters for Kazal inhibitor and benzamidine association to the binary peptide/zymogen adducts are more favorable than those observed for ligand binding to the proenzyme alone, although never as much as those reported for the formation of bovine beta-trypsin/Kazal inhibitor and bovine beta-trypsin/benzamidine adducts. Analogously, the affinity of activating peptides for the binary proenzyme/Kazal inhibitor and binary proenzyme/benzamidine complexes is higher than that observed for peptide binding to free bovine trypsinogen. Differences in affinity for ligand binding to free bovine trypsinogen, to its binary adducts and to bovine beta-trypsin suggest the presence of different activation levels of the proenzyme, none of which structurally coincide with that achieved in bovine beta-trypsin. The existence of different discrete states suggests that the zymogen-to-active enzyme transition should not be considered as a two-state process but as a multistep event.
Ascenzi
P
P
Department of Biochemical Sciences, University of Rome 'La Sapienza', Italy.
Coletta
M
M
Amiconi
G
G
Bolognesi
M
M
Guarneri
M
M
Menegatti
E
E
D000578:Amidines / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001550:Benzamidines / Q000378:metabolism*; D002417:Cattle; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000502:physiology*; D011485:Protein Binding; D014357:Trypsin / Q000502:physiology*; D014359:Trypsin Inhibitor, Kazal Pancreatic / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000378:metabolism*; D014362:Trypsinogen / Q000378:metabolism*
D000578:Amidines; D001550:Benzamidines; D008024:Ligands; D010455:Peptides; D014361:Trypsin Inhibitors; D014359:Trypsin Inhibitor, Kazal Pancreatic; D014362:Trypsinogen; D014357:Trypsin