Hydrogen ion uptake by chymotrypsin during reversible binding of specific substrate is shown to be due to an ionizing group of the enzyme with a pK(apparent) approximately 9 in the free enzyme. This pK(apparent) is shifted to higher value in the enzyme-substrate complexes. Previous results indicating an equilibrium, controlled by this ionizing group, between active and inactive conformational forms of chymotrypsin are confirmed.

tained are presented in Table II. The analysis of two selected naturally occurring minerals is presented in Table III.The usual precautions must be taken when working with trace elements in the ranges studied. In addition, the glassware should be cleaned with hot nitric acid followed by ammonium hydroxide before rinsing with distilled water. Beakers should be used which have not been subjected to chromic acid cleaning solution. Standard solutions, especially Ag, should be prepared fresh from concentrated stock solutions, as the metals tend to be rapidly adsorbed on the walls of the containers. Samples ground in a tungsten carbide mixing vial had cobalt contamination.The recovery of Mn was not quantitative although precipitation was carried out at a pH of 5.9. Sources of Mn loss result from improper pH of precipitation and loss in the digestion process.

Chemical relaxation experiments were conducted on the reaction of α-chymotrypsin, with the competitive inhibitor proflavin and the substrate analogue TAME (tosylarginine methyl ester) in phosphate buffer, pH 6.7, observing transmission changes at 465 mμ. Two chemical relaxation processes were observed with the slow one attributed to a monomolecular interconversion of the enzyme–substrate complex. The concentration dependence of the reciprocal fast relaxation time constant only agrees with the equations derived for the involvement of a labile ternary complex between enzyme, substrate, and inhibitor (as simplest model).