Binding of Competitive Inhibitors to S-Chymotrypsin in the Alkaline pH Region. Competitive Inhibition Kinetics and Proton-Uptake Measurements

Pablo Valenzuela, Myron L. Bender

Resultado de la investigación: Contribución a una revistaArtículo

33 Citas (Scopus)

Resumen

The binding of the three competitive inhibitors benzyl alcohol, tryptophol, and N-acetyl-D-tryptophanamide to α- and δ-chymotrypsins was studied over the pH range 7 to 11 by competitive inhibition kinetics using N-furyl-acryloyl-L-tryptophan methyl ester as substrate. The results indicate that the binding of these inhibitors to δ-chymotrypsin exhibits a pH dependence significantly different from the pH dependence obtained with α-chymotrypsin. Analysis of K, vs. pH profiles for the interaction of benzyl alcohol, tryptophol, and N-acetyl-D-tryptophanamide with δ-chymotrypsin indicates that the pKa of an ionizing group of the enzyme (9.2, 9.5, and 9.2, respectively) is shifted to a pKa of 10.0, 10.1, and 9.8, respectively, in the enzyme-inhibitor complex. This behavior differs from that of α-chymotrypsin, where, in agreement with previous reports, the binding of the three inhibitors was found to be strictly dependent on the ionization of a group in the enzyme with a pKa of 9.0 that apparently shifts upon binding of inhibitor to a value of 11.5 or higher. The proton uptake that accompanies the binding of the three inhibitors to δ-chymotrypsin was also measured as a function of pH. A good agreement between the measured proton uptake and the predicted values from the corresponding pKa shifts was found. It is concluded that the different behavior of α- and δ-chymotrypsins at high pH reflects a difference in the binding abilities of the enzymes. The present hypotheses regarding the conformation and activity of α-chymotrypsin at alkaline pH are discussed in the light of this particular behavior of δ-chymotrypsin.

Idioma originalInglés
Páginas (desde-hasta)2440-2446
Número de páginas7
PublicaciónBiochemistry
Volumen9
N.º12
DOI
EstadoPublicada - 1 jun 1970

Áreas temáticas de ASJC Scopus

  • Bioquímica

Huella Profundice en los temas de investigación de 'Binding of Competitive Inhibitors to S-Chymotrypsin in the Alkaline pH Region. Competitive Inhibition Kinetics and Proton-Uptake Measurements'. En conjunto forman una huella única.

  • Citar esto