Differential scanning calorimetric study of carboxypeptidase B, procarboxypeptidase B and its globular activation domain.
High-sensitivity differential scanning calorimetry has been applied to the study of porcine pancreatic carboxypeptidase B, the proenzyme and its 81-residue activation domain. The thermal study has been carried out over a range of scan rates, ionic strengths and pH values. The thermal unfolding of the isolated activation domain has been found to be reversible and corresponds to that of a typical compact globular structure, with melting temperatures higher than those of the enzyme and proenzyme. Both proteins, on the other hand, undergo an irreversible, highly scan-rate-dependent thermal denaturation under all the experimental conditions investigated. The denaturation of the enzyme at pH 7.5 and the proenzyme at pH 7.5 and 9.0 follows the two-state irreversible model [Sánchez-Ruiz, J.M., López-Lacomba, J.L., Cortijo, M. & Mateo, P.L. (1988) Biochemistry 27, 1648-1652]. Thus the kinetic constants and activation parameters of the denaturation process could be obtained and compared to those for other proteins, particularly those of the closely related carboxypeptidase A system.
Conejero-Lara
F
F
Departamento de Química Física (Facultad de Ciencias), Universidad de Granada, Spain.
Sánchez-Ruiz
J M
JM
Mateo
P L
PL
Burgos
F J
FJ
Vendrell
J
J
Avilés
F X
FX
D000818:Animals; D002152:Calorimetry, Differential Scanning; D043424:Carboxypeptidase B; D002268:Carboxypeptidases / Q000737:chemistry*; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000737:chemistry*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D009994:Osmolar Concentration; D010179:Pancreas / Q000201:enzymology*; D011487:Protein Conformation; D011489:Protein Denaturation; D013552:Swine; D013816:Thermodynamics
D004792:Enzyme Precursors; D002268:Carboxypeptidases; D043424:Carboxypeptidase B