Abstract
Purpose :
Lecithin retinol acyltransferase (LRAT) is responsible for transforming all-trans retinol into all-trans retinyl ester in the visual cycle. Our preliminary nuclear magnetic resonance data of a truncated form of LRAT (residues 30-196) suggests the presence of an amphiphilic segment (residues 96-107) which could bind membranes. In addition, other authors have postulated that a segment near this amphiphilic segment (residues 71-103), which is found uniquely in LRAT, could be involved in its membrane binding. However, no evidence was shown to support their hypothesis. The objectives of this study are therefore to determine the secondary structure and the membrane binding of these segments of LRAT.
Methods :
The amphiphilic segment of LRAT was synthesized commercially. In addition, the human and murine forms of the unique segment were synthesized by the Plateforme de Chimie Médicinale du Centre de Recherche du CHU de Québec. The secondary structure of these peptides was determined by circular dichroism (CD) and their membrane binding was studied using Langmuir monolayers.
Results :
The CD spectra show that all segments have a predominantly α-helical structure in methanol. However, in an aqueous medium, the secondary structure of the amphiphilic segment and the murine unique segment is mostly in random coil, whereas the human unique segment is insoluble. All segments bind strongly to monolayers of different types of phospholipids. Indeed, maximum insertion pressure (MIP) values of 44 ± 2, 47 ± 3 and 57 ± 5 mN/m were obtained for the amphiphilic segment in the presence of dioleoylphosphocholine, dioleoylphosphoethanolamine and distearoylphoshocholine, respectively. MIP values of 36 ± 2, 40 ± 1, 35 ± 1 and 40 ± 2 mN/m were obtained for the murine unique segment in the presence of dioléoylphosphocholine, dioléoylphosphoéthanolamine, palmitoyl-arachidonoyl phosphocholine and palmitoyl-arachidonoyl phosphoethanolamine, respectively.
Conclusions :
The amphiphilic segment and the murine unique segment both have a high affinity for membranes since all the MIP values obtained are superior to the estimated lateral pressure of membranes (30 mN/m). This suggests that these segments could be involved in the membrane binding of LRAT. This study gives a better understanding of the role of these segments in the function of LRAT. Langmuir monolayer measurements of the human unique segment are in progress.
This is a 2021 ARVO Annual Meeting abstract.