Studies in Bile-Salt Solutions. XXII. The Effect of Reversed Micelles and of Aerosol-OT Aqueous Micelles on the Esterase Activity of Bile-Salt-Stimulated Human-Milk Lipase. Determination of Enzyme-Inhibitor Complex Dissociation Constants

Abstract

The esterase activity of bile-salt-stimulated human milk lipase has been measured against several 4-nitrophenyl alkanoate esters, including 4-nitrophenyl propionate, in reversed micellar solutions of several surfactants. The reaction media used were Aerosol-OT in isooctane, hexadecyltrimethylammonium bromide in chloroform/n-octane (1 : 1 v/v) or in chloroform, Triton X-100 in carbon tetrachloride or isooctane, lecithin in n-octane or diethyl ether/methanol (95 : 5 v/v) or benzene, and Brij 56 in cyclohexane . The reaction conditions varied over a range of cosolubilized water concentrations and initial pH values of added borate, glycine and Tris buffer solutions. In all cases decreased enzymic activity was observed, even if sodium taurocholate was present in the reaction medium. A detailed examination has been made on the effect of changing concentrations of 4-nitrophenyl propionate and of Aerosol-OT on the esterase activity of bile-salt-stimulated human milk lipase at 298 K in aqueous solutions of Tris buffer at pH 7.5. The inhibition induced by Aerosol-OT was identified as being reversible and mixed. The dissociation constants of the enzyme-surfactant complex and of the (enzyme-ester)-surfactant complex have been calculated to be equal to 0.125 and 0.48 mmol 1^-1, respectively.