title: How gastric lipase, an interfacial enzyme with a Ser-His-Asp catalytic triad, acts optimally at acidic pH.
abstract: Gastric lipase is active under acidic conditions and shows optimum activity on insoluble triglycerides at pH 4. The present results show that gastric lipase also acts in solution on vinyl butyrate, with an optimum activity above pH 7, which suggests that gastric lipase is able to hydrolyze ester bonds via the classical mechanism of serine hydrolases. These results support previous structural studies in which the catalytic triad of gastric lipase was reported to show no specific features. The optimum activity of gastric lipase shifted toward lower pH values, however, when the vinyl butyrate concentration was greater than the solubility limit. Experiments performed with long-chain triglycerides showed that gastric lipase binds optimally to the oil-water interface at low pH values. To study the effects of the pH on the adsorption step independently from substrate hydrolysis, gastric lipase adsorption on solid hydrophobic surfaces was monitored by total internal reflection fluorescence (TIRF), as well as using a quartz crystal microbalance. Both techniques showed a pH-dependent reversible gastric lipase adsorption process, which was optimum at pH 5 (Kd = 6.5 nM). Lipase adsorption and desorption constants (ka = 147,860 M(-1) s(-1) and kd = 139 x 10(-4) s(-1) at pH 6) were estimated from TIRF experiments. These results indicate that the optimum activity of gastric lipase at acidic pH is only "apparent" and results from the fact that lipase adsorption at lipid-water interfaces is the pH-dependent limiting step in the overall process of insoluble substrate hydrolysis. This specific kinetic feature of interfacial enzymology should be taken into account when studying any soluble enzyme acting on an insoluble substrate.
authors.affiliation: Laboratoire d'Enzymologie Interfaciale et de Physiologie de la Lipolyse, CNRS UPR 9025, IBSM, Marseille, France.
authors.forename: Henri
authors.initials: H
authors.lastname: Chahinian
authors.forename: Torben
authors.initials: T
authors.lastname: Snabe
authors.forename: Coralie
authors.initials: C
authors.lastname: Attias
authors.forename: Peter
authors.initials: P
authors.lastname: Fojan
authors.forename: Steffen B
authors.initials: SB
authors.lastname: Petersen
authors.forename: Frédéric
authors.initials: F
authors.lastname: Carrière
mesh_terms: D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism; D004285:Dogs; D006639:Histidine / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000737:chemistry* / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D013270:Stomach / Q000201:enzymology*; D003313:Zea mays / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D001224:Aspartic Acid; D012694:Serine; D006639:Histidine; D008049:Lipase