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LCAT is an enzyme responsible for the formation of cholesteryl esters from unesterified cholesterol (UC) and phospholipid (PL) molecules in HDL particles. However, it is poorly understood how LCAT interacts with lipoproteins and how apoA-I activates it. Here we have studied the interactions between LCAT and lipids through molecular simulations. In addition, we studied the binding of LCAT to apoA-I-derived peptides, and their effect on LCAT lipid association-utilizing experiments. Results show that LCAT anchors itself to lipoprotein surfaces by utilizing nonpolar amino acids located in the membrane-binding domain and the active site tunnel opening. Meanwhile, the membrane-anchoring hydrophobic amino acids attract cholesterol molecules next to them. The results also highlight the role of the lid-loop in the lipid binding and conformation of LCAT with respect to the lipid surface. The apoA-I-derived peptides from the LCAT-activating region bind to LCAT and promote its lipid surface interactions, although some of these peptides do not bind lipids individually. The transfer free-energy of PL from the lipid bilayer into the active site is consistent with the activation energy of LCAT. Furthermore, the entry of UC molecules into the active site becomes highly favorable by the acylation of SER181.

Age related macular degeneration (AMD) is a progressive, neurodegenerative disorder that leads to the severe loss of central vision in elderlies. The health of retinal pigment epithelial (RPE) cells is critical for the onset of AMD. Chronic oxidative stress along with loss of lysosomal activity is a major cause for RPE cell death during AMD. Hence, development of a molecule for targeted lysosomal delivery of therapeutic protein/drugs in RPE cells is important to prevent RPE cell death during AMD. Using human RPE cell line (ARPE-19 cells) as a study model, we confirmed that hydrogen peroxide (H2O2) induced oxidative stress results in CD44 cell surface receptor overexpression in RPE cells; hence, an important target for specific delivery to RPE cells during oxidative stress. We also demonstrate that the known nucleic acid CD44 aptamer - conjugated with a fluorescent probe (FITC) - is delivered into the lysosomes of CD44 expressing ARPE-19 cells. Hence, as a proof of concept, we demonstrate that CD44 aptamer may be used for lysosomal delivery of cargo to RPE cells under oxidative stress, similar to AMD condition. Since oxidative stress may induce wet and dry AMD, both, along with proliferative vitreoretinopathy, CD44 aptamer may be applicable as a carrier for targeted lysosomal delivery of therapeutic cargoes in ocular diseases showing oxidative stress in RPE cells.