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.

Boosting of retinol activity using novel lecithin: Retinol acyltransferase inhibitors

“…Interestingly, the acyl chain length of our LRAT inhibitors affected their inhibitory activity with the shorter variants being more active in accordance with the substrate specificity reported by Horchani et al [29]. Using molecular modelling, the above-described SAR could be investigated in a structural setting.…”

ATGL and CGI-58 are lipid droplet proteins of the hepatic stellate cell line HSC-T6

“…Similarly, LRAT exhibits virtually no preference for neither FA16:0 nor FA18:1, although it exhibits a clear preference for short acyl chains ( e.g. for FA7:0 ~50-fold higher activity) (55). Furthermore, this flexibility indicates that also the neutral lipid store of HSCs quickly responds to FA and ROH overload and channels excess lipids for storage.…”

Hepatic stellate cells retain the capacity to synthesize retinyl esters and to store neutral lipids in small lipid droplets in the absence of LRAT

“…Lecithin:retinol acyltransferase (LRAT) catalyzes a trans-esterification reaction that occurs between retinol and phosphatidylcholine (PC), transferring the fatty acid at the sn-1 position of PC to retinol [9][10][11]. LRAT does not have a clear preference for physiologically occurring fatty acids at the sn-1 position of PC [12]. The other enzymatic activity catalyzes an acyl coenzyme A:retinol acyltransferase (ARAT) reaction.…”