Mechanism of pancreatic lipase action. 2. Catalytic properties of modified lipases

Abstract

Reaction of lipase [EC 3.1.1.3] [pig] with diethyl pyrocarbonate results in the modification of 3 histidine residues. One is highly reactive, although without affecting the activity, while the 2 others react more slowly with a concomitant loss of activity on both dissolved and emulsified substrates. As previously shown, lipase can also be modified either by reaction of 5 carboxyl groups with carbodiimide (5N-lipase) or by esterification of 1 serine residue with diethyl p-nitrophenyl phosphate (DP-lipase). In the 3 cases the activity on emulsified substrates is abolished. The modification of histidine residues results also in a loss of activity on dissolved substrates, suggesting that the essential histidine is at (or close to) the active site. The ability of lipase to be adsorbed on siliconized glass beads is not impaired in this reaction. By contrast, 5N-lipase is still able to hydrolyze dissolved monomeric substrates and to adsorb on siliconized glass beads. Therefore, the essential carboxyl group is assumed to play an important role in the interfacial activation. Finally, since DP-lipase is still fully active on dissolved p-nitrophenyl acetate, the serine residue, which was suggested to be the acylable one, is more likely implicated in the recognition and the binding to interfaces, as confirmed by the inability of DP-lipase to be adsorbed on siliconized glass beads.