title: Enzymatic activity of Lecithin:retinol acyltransferase: a thermostable and highly active enzyme with a likely mode of interfacial activation.
abstract: Lecithin:retinol acyltransferase (LRAT) plays a major role in the vertebrate visual cycle. Indeed, it is responsible for the esterification of all-trans retinol into all-trans retinyl esters, which can then be stored in microsomes or further metabolized to produce the chromophore of rhodopsin. In the present study, a detailed characterization of the enzymatic properties of truncated LRAT (tLRAT) has been achieved using in vitro assay conditions. A much larger tLRAT activity has been obtained compared to previous reports and to an enzyme with a similar activity. In addition, tLRAT is able to hydrolyze phospholipids bearing different chain lengths with a preference for micellar aggregated substrates. It therefore presents an interfacial activation property, which is typical of classical phospholipases. Furthermore, given that stability is a very important quality of an enzyme, the influence of different parameters on the activity and stability of tLRAT has thus been studied in detail. For example, storage buffer has a strong effect on tLRAT activity and high enzyme stability has been observed at room temperature. The thermostability of tLRAT has also been investigated using circular dichroism and infrared spectroscopy. A decrease in the activity of tLRAT was observed beyond 70°C, accompanied by a modification of its secondary structure, i.e. a decrease of its α-helical content and the appearance of unordered structures and aggregated β-sheets. Nevertheless, residual activity could still be observed after heating tLRAT up to 100°C. The results of this study highly improved our understanding of this enzyme.
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Habib
authors.initials: H
authors.lastname: Horchani
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Sylvain
authors.initials: S
authors.lastname: Bussières
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Line
authors.initials: L
authors.lastname: Cantin
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Mustapha
authors.initials: M
authors.lastname: Lhor
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Jean-Sébastien
authors.initials: JS
authors.lastname: Laliberté-Gemme
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada.
authors.forename: Rock
authors.initials: R
authors.lastname: Breton
authors.affiliation: CUO-Recherche, Hôpital du Saint-Sacrement, Centre de Recherche du CHU de Québec, Québec, Québec, Canada; Département d'Ophtalmologie, Faculté de Médecine, Université Laval, Québec, Québec, Canada; Regroupement Stratégique PROTEO, Université Laval, Québec, Québec, Canada. Electronic address: christian.salesse@fmed.ulaval.ca.
authors.forename: Christian
authors.initials: C
authors.lastname: Salesse
mesh_terms: D000217:Acyltransferases / Q000737:chemistry* / Q000235:genetics; D002942:Circular Dichroism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D007700:Kinetics; D054709:Lecithins / Q000737:chemistry*; D008823:Micelles; D010742:Phospholipid Ethers / Q000737:chemistry*; D055550:Protein Stability; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D013055:Spectrophotometry, Infrared; D013379:Substrate Specificity; D014801:Vitamin A / Q000737:chemistry*
chemical_list: C052298:1,2-dihexadecyl-sn-glycero-3-phosphocholine; D054709:Lecithins; D008823:Micelles; D010742:Phospholipid Ethers; D011994:Recombinant Proteins; D014801:Vitamin A; D000217:Acyltransferases; C056942:lecithin-retinol acyltransferase
keywords: Enzymatic activity; Lecithin:retinol acyltransferase; Retinol; Thermostability; Truncated LRAT; Visual cycle