PANCREATIC LIPASE/COLIPASE BINDING SITE INVESTIGATION

Abstract

Pancreatic lipase is responsible for fat digestion in the intestine. The enzyme, which belongs to a special class of esterases, realizes an heterogeneous catalysis. In vivo, because of the strong inhibitory effect of bile salt,the lipase action requires the presence of a small pancreatic protein, colipase, the function of which is to anchor lipase to the bile salt coated lipid interface. The function of the lipase/colipase system requires the presence of two topographically distinct binding sites on both proteins, an interfacial binding site and a protein binding site. Up to now, only the colipase interfacial binding site is well documented. In order to locate the lipase/colipase binding site on each partner, a covalent cross-linked complex has been obtained using carbodiimides. Immunological analysis of the complex clearly confirms the presence of lipase and colipase. The complex has a Mr (60 kDa) consistent with a stoichiometry of one mol colipase per mol lipase and retains its catalytic efficiency towards emulsified substrates. Moreover, the use of carbodiimides to cross-link lipase and colipase unambiguously shows the participation of ion-pairing in the interaction between the two proteins.