Biochemical Characterization and Molecular Modeling of Pancreatic Lipase from a Cartilaginous Fish, the Common Stingray (Dasyatis pastinaca)

Abstract

In order to identify fish enzymes displaying novel biochemical properties, we have chosen the common stingray (Dasyatis pastinaca), one of the most primitive living jawed aquatic vertebrates as a starting biological material to purify a lipase. A stingray pancreatic lipase (SPL) was purified from delipidated pancreatic powder. The SPL molecular weight was around 55 kDa which is slightly higher than that of known classical pancreatic lipases (50 kDa). This increase in the molecular weight was due to glycosylation. Like classic pancreatic lipases, SPL was found to be much more active on short-chain triacylglycerols than on long-chain ones. Natural detergents act as inhibitors of the SPL activity. This inhibition can be reversed by the addition of stingray colipase. Starting from total pancreatic messenger RNAs (mRNAs), partial stingray pancreatic lipase complementary DNA (cDNA) was synthesized by reverse transcriptase-polymerase chain reaction (RT-PCR) and cloned into the PGEM-T vector. Partial amino acid sequence of the SPL was homologous to that of Japanese eel, porcine, and human pancreatic lipases. A 3D structure model of the sequenced part of SPL was built using the 3D structure of porcine pancreatic lipase as template, since both lipases shared an amino acid sequence identity of 60%.

Biochemical characterization, cloning and molecular modeling of a digestive lipase from red seabream (Pagrus major): Structural explanation of the interaction deficiency with colipase and lipidic interface

“…Similar results were previously obtained with purified lipase from the annular seabream and other digestive lipases isolated from sardine and crab . Furthermore, this behavior is different from that of stingray pancreatic lipase, which showed an absolute requirement for bile salts .…”

“…The optimal activity of the purified lipase was recorded at pH 9 and at 50°C. Thus, the RsDL, like AsDL , is more thermoactive than most of fish digestive lipases, previously characterized, which display a maximal activity around 40°C . The optimum pH value for the RsDL activity was close to that reported for digestive lipolytic enzymes from other fish species such as sardine and golden grey mullet .…”

Dissecting the Interaction Deficiency of a Cartilaginous Fish Digestive Lipase with Pancreatic Colipase: Biochemical and Structural Insights

“…The deduced polypeptide sequences of SmDL and European eel digestive lipases comprise 451 and 452 amino acids, respectively. High amino acid sequence identities were found between SmDL and other cartilaginous fish lipases like Callorhinchus milii (68%) and Dasyatis pastinaca (79%) [36]. The EeDL amino acid sequence displayed prominent similarity with those of other fish species like Anguilla japonica (97%) and red seabream (63%).…”

“…The complete sequences of SmDL and EeDL ( Figure S2) consisting of 1.35 kb were submitted in the GenBank and were assigned the accession numbers KY548033 and KY548034, respectively. The nucleotide BLAST analysis for SmDL showed that this enzyme shared high identity (77%-87%) with pancreatic lipase genes from other cartilaginous fish: 87% with the whale shark (Rhincodon typus), 82% with the common stingray (Dasyatis pastinaca) [36], and 77% with the elephant shark (Callorhinchus milii). The EeDL ( Figure S2B) shared high identities of 96% and 75% with the Japanese eel (Anguilla japonica) and hareng (Clupea harengus) lipase genes, respectively.…”

Integrative computational approach for genome-based study of microbial lipid-degrading enzymes

“…In addition, 3D structural analysis methods have been applied to lipid-degrading enzymes in various organisms, including humans and animals (Fendri et al 2006;Lampidonis et al 2008;Cornaciu et al 2011;Ataya 2012;Bouchaâla et al 2015;Ganesan et al 2015;Mahankali et al 2015). Specifically, structural studies of lipid-degrading enzymes in microbes have been reported (Vasel et al 1993;Gaskin et al 2001;Mandrich et al 2004;Jan et al 2015).…”