Aspects on the catalysis of lipase from porcine pancreas (type VI-s) in aqueous media: development of ion-pairs

Abstract

This article reports a first contribution for the elucidation of catalytic mechanism of Lipase from porcine pancreas, type VI-s (PPL), in hydrolyzing an ester substrate in aqueous media. The conclusions were based on the pH-profiles of Michaelis-Menten parameters k cat/Km, k cat and Km, as well as on the absolute temperature profile of k cat/Km, obtained during the hydrolysis of p-nitrophenyl laurate by PPL. It was found that (a) PPL performs catalysis by means of ion pairs formed either as Ser152-Ο-/His263-Im+H and/or Carbonyl-Ο-/His263-Im+H, (b) the parameter k cat/Km equals to k1 and thus ES is formed and destroyed in the course of a series of consecutive reactions governed by the dynamic constant K S = k2/k1, and (c) the hydrolysis of substrate is assisted by a hydrogen bond developed between deprotonated Asp176 and the positively charged imidazole of His263 across a pKa-value 3.85, necessary for efficient catalysis

Fungal endophytes associated with Viola odorata Linn. as bioresource for pancreatic lipase inhibitors

“…The human PL (EC 3.1.1.3) is a primary digestive enzyme secreted from the exocrine glands of pancreas and is primarily involved in the hydrolysis of ester bonds of triglycerides [ 21 ]. The hydrolysis of the triglyceride esters by PL is represented by a series of events, initiated by an interfacial activation by the hydrophobic alkyl chains of the triglycerides, resulting in the open lid conformation followed by the nucleophilic attack of Ser 152 on to the carbonyl carbon of the ester linkage of the triglycerides [ 22 , 23 ].…”

Molecular, biochemical and kinetic analysis of a novel, thermostable lipase (LipSm) from Stenotrophomonas maltophilia Psi-1, the first member of a new bacterial lipase family (XVIII)

“…All activity and kinetic measurements throughout this study were carried out in aqueous buffers of 100 mM Tris–HCl, pH 8.0 containing 1% Triton X-100 w/v at 40 °C, except, if otherwise stated. In all cases, the aqueous reaction media contained 5% v/v dimethylsulfoxide (DMSO), to increase the solubility of the used substrates [ 6 , 23 , 24 , 46 ].…”

“…esterifications or transesterifications [ 2 , 3 ]. Due to these unique properties, lipases are enzymes of considerable biotechnological interest and find use in a broad spectrum of applications [ 4 ] including, among others, food technology, bioremediation, chemical industry and medical sciences [ 2 , 5 , 6 ].…”

Effective Kinetic Methods and Tools in Investigating the Mechanism of Action of Specific Hydrolases

“…Mechanistic features, similar to those of serine proteases, have been reported also for lipases, as it is the oxyanion hole, although its development differs in these enzymes due to their structural particularities (Aloulou et al, 2006). Recently, a full mechanism of action for the lipase from bovine pancreas (PPL) has been reported and it is analogous to this reported for serine proteases (figure 4a) (Kokkinou et al, 2011). However, it is essential to show that the mechanism of fatty acid ester hydrolysis by lipases in micelles, small aggregates or emulsion particles is noticeably different.…”