Enzyme-substrate interaction in lipid monolayers. I. Experimental conditions and fundamental kinetics.

A systematic study has shown the importance of the different factors which are concerned with the action of lipase on a substrate (1,3-didecanoylglycerol). These consist of a) the process of adsorption of lipase to the surface, b) the necessity of limited stirring to reach equilibrium, and c) the persistence during the reaction process of the enzyme molecules adsorbed on the monolayer. On the basis of this preliminary investigation, a technique was established to analyze the mechanism of lipase action with defined quantities of enzyme and lipid segregated in the monolayer. Thus, the process of the reaction itself is separated from the adsorption process, and it is demonstrated that the quantity of substrate hydrolyzed per minute depends only on the quantity of initially adsorbed lipase and not on the quantity of substrate or on the surface concentration of the enzyme. An appropriate new definition of the rate is consequently adopted.