Influence of the Lipid Moiety Structure on the Insertion/Release of Glycolipids in/from the Cell: A Study with Synthetic Analogs.

Many important aspects of intercellular glycosphingolipid (GSL) transport remain unknown, in particular how GSLs cross the glycocalyx of the cell-donor and cell-acceptor. The use of synthetic analogs instead of natural GSLs alleviates the study. However, in order to obtain adequate results, we must know how simplifications in the structure of a synthetic probe affect its properties of interest to us. In particular, how the replacement of a ceramide residue in GSL with a synthetic analog affects the release and insertion kinetics. To this end, we compared the properties of synthetic analogs, one of which contains natural ceramide moiety, while two others have lipid residues that are more convenient in terms of the probes chemical synthesis. FSL (Function-Spacer-Lipid) constructs containing DOPE (1,2-O-dioleoyl-sn-glycero-3-phospho ethanolamine), cholesterol (Chol), or C18-ceramide (Cer) lipid fragments (L) were compared. In all three probes, the lipid residues were linked to biotin with the hydrophilizing CMG2-spacer-arm. Using confocal microscopy, the similarity of the localization patterns of these three probes, both with each other and with the previously studied glycoprobes (F = glycan), was shown, namely: they were observed in the form of patches of an average size of 0.3 μm but were not detected at all in cell rafts. At the same time, the following differences were observed in the properties of the three studied FSLs: (1) the process of insertion of the ceramide construct reaches a plateau within three hours, while the other two within one hour; (2) the amount of inserted ceramide construct is less than the other two; and (3) release of the cholesterol construct from the cells where it is preinserted is slower than the other two. Thus, despite some quantitative differences, qualitatively all three constructs behave similarly, that is, they are interchangeable when studying the incorporation/release of the FSL into/from the cell. [ABSTRACT FROM AUTHOR]