Inhibition of lipases by -polylysine

Oral administration of -polylysine to rats reduced the peak plasma triacylglycerol concentration. In vitro, -polylysine and polylysine strongly inhibited the hydrolysis, by either pancreatic lipase or carboxylester lipase, of trioleoylglycerol (TO) emulsified with phosphatidylcholine (PC) and taurocholate. The -polylysine concentration required for complete inhibition of pancreatic lipase, 10 g/ml, is 1,000 times lower than that of BSA required for the same effect. Inhibition requires the presence of bile salt and, unlike inhibition of lipase by other proteins, is not reversed by supramicellar concentrations of bile salt. Inhibition increases with the degree of polylysine polymerization, is independent of lipase concentration, is independent of pH between 5.0 and 9.5, and is accompanied by an inhibition of lipase binding to TO-PC emulsion particles. However, -polylysine did not inhibit the hydrolysis by pancreatic lipase of TO emulsions prepared using anionic surfactants, TO hydrolysis catalyzed by lingual lipase, or the hydrolysis of a water-soluble substrate. In the presence of taurocholate, -polylysine becomes surface active and adsorbs to TO-PC monomolecular films. These results are consistent with -polylysine and taurocholate forming a surface-active complex that binds to emulsion particles, thereby retarding lipase adsorption and triacylglycerol hydrolysis both in vivo and in vitro. —Tsujita, T., M. Sumiyoshi, T. Takaku, W. E. Momsen, M. E. Lowe, and H. L. Brockman. Inhibition of lipases by -polylysine. J. Lipid Res. 2003. 44: 2278–2286. Supplementary key words basic peptide • complex • lipid emulsion • lipid monolayer In mammals, dietary neutral lipid digestion is commonly assumed to be mediated by three main enzymes: preduodenal (lingual or gastric) lipase, carboxylester lipase (cholesterol esterase), and pancreatic lipase (1). Typical substrates for these enzymes are water-insoluble longchain triacylglycerols. In contrast to their lipid substrates, these lipases are water soluble. Thus, for catalysis to occur, these enzymes must be adsorbed to the lipid surfaces and, therefore, the quality of the surface of substrate lipids is an important factor for lipase activity. Enzymes are sometimes activated or denatured by surface adsorption (2). As a consequence of these properties, amphiphilic substances, acting as emulsifiers, would be expected to influence the lipase reaction rate. It is well known that bile salts and synthetic detergents behave as inhibitors of lipolysis (3, 4). Amphiphilic proteins, such as BSA and -lactoglobulin, have been shown to inhibit lipase activity toward its triglyceride substrate (5, 6). Previously, we demonstrated that a basic protein, protamine, strongly inhibited the hydrolysis of trioleoylglycerol (TO) emulsified with phosphatidylcholine (PC) (7). Polylysine, another basic protein, has been suggested as a food antiseptic. Polylysine binds to some proteins, nucleic acids, viruses, or bacteria through electrostatic or hydrophobic interaction and inhibits their functions. Another form of polylysine, -polylysine, is synthesized by linking the -carboxyl groups of lysine with its -amino groups. As a consequence of this linkage, it is not hydrolyzed by proteases, such as trypsin, but retains its basic character. Therefore, -polylysine is a candidate for an agent that inhibits intestinal lipid absorption while resisting proteolysis. The experiments presented in this report were undertaken to evaluate this possibility and elucidate its mode of action. MATERIALS AND METHODS