Binding of the soybean Bowman-Birk proteinase inhibitor and of its chymotrypsin and trypsin inhibiting fragments to bovine alpha-chymotrypsin and bovine beta-trypsin. A thermodynamic study.
The effect of pH and temperature on the apparent association equilibrium constant (Ka) for the binding of the soybean Bowman-Birk proteinase inhibitor (BBI) and of its chymotrypsin and trypsin inhibiting fragments (F-C(p), F-T(p) and F-T(t), respectively) to bovine alpha-chymotrypsin (alpha-chymotrypsin) and bovine beta-trypsin (beta-trypsin) has been investigated. On the basis of Ka values, the proteinase inhibitor affinity can be arranged as follows: alpha-chymotrypsin: BBI approximately beta-trypsin:BBI approximately beta-trypsin:F-T(t) approximately beta-trypsin:F-T(p) much greater than alpha-chymotrypsin:F-C(p). F-C(p), F-T(p) and F-T(t) do not inhibit beta-trypsin and alpha-chymotrypsin action, respectively. On lowering the pH from 9.5 to 4.5, values of Ka for BBI, F-C(p), F-T(p) and/or F-T(t) binding to alpha-chymotrypsin and beta-trypsin decrease, thus reflecting the acid-pK shift of the invariant His57 catalytic residue from 7.0, in the free enzymes, to 5.2, in the proteinase:inhibitor complexes. Considering the known molecular models, the observed binding behaviour of BBI, F-C(p), F-T(p) and F-T(t) was related to the inferred stereochemistry of the proteinase:inhibitor contact regions.
Ascenzi
P
P
Department of Biochemical Sciences, University of Rome La Sapienza, Italy.
Amiconi
G
G
Bolognesi
M
M
Menegatti
E
E
Guarneri
M
M
D000818:Animals; D002417:Cattle; D002918:Chymotrypsin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D013816:Thermodynamics; D014357:Trypsin / Q000378:metabolism*; D014358:Trypsin Inhibitor, Bowman-Birk Soybean / Q000378:metabolism*
D010446:Peptide Fragments; D014358:Trypsin Inhibitor, Bowman-Birk Soybean; D002918:Chymotrypsin; D014357:Trypsin