PRODUCT INHIBITION OF ALPHA-CHYMOTRYPSIN IN REVERSE MICELLES

The alpha-chymotrypsin-catalyzed hydrolysis of succinyl-L-alanyl-L-alanyl-L-prolyl-L-phenylalanyl p-nitroanilide has been studied in reverse micelles of sodium bis(2-ethylhexyl)sulfosuccinate (AOT) in isooctane. It has been found that alpha-chymotrypsin is strongly inhibited competitively by the acidic peptide product which is formed during the course of the reaction. It has also been shown that the application of the integrated form of the Michaelis-Menten equation can be useful to detect possible inhibition effects and abnormal kinetic behavior of enzymes in reverse micelles. Furthermore, it has been shown that the turnover number (k(cat)) at low water content is lower than in water and increases as the water content in the system (w(o) = [H2O]/[AOT]) increases, k(cat) reaching the value in water at high w(o). If however, initial velocity data, as obtained under conditions where the enzyme is not saturated with substrate, are plotted against w(o), the curves are bell-shaped, with a maximum around w(o) = 15.