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  • Title: Behaviour of complex oligosaccharides at a bilayer membrane surface: probed by 2H-NMR.
    Author: Barber KR, Hamilton KS, Rigby AC, Grant CW.
    Journal: Biochim Biophys Acta; 1994 Mar 23; 1190(2):376-84. PubMed ID: 8142439.
    Abstract:
    Deuterium wideline NMR was used in an attempt to directly assess oligosaccharide arrangement and motional characteristics of complex glycosphingolipids dispersed as minor components in phospholipid membranes. A convenient, general synthetic approach was developed which involved replacement of the acetate group of amido sugars with deuteroacetate (-COCD3). This provided excellent signal-to-noise when applied to the terminal GalNAc residue of globoside, and the terminal NANA residue of GM1. Simultaneously, globoside and GM1 fatty acids were replaced with stearic acid deuterated at C-2- a probe location sensitive to glycolipid hydrophobic backbone orientation and rigid body motion. Deuterated GM1 and globoside were studied by 2H-NMR in bilayers of 1-palmitoyl-2-oleoyl phosphatidylcholine, in the presence and absence of physiological quantities of cholesterol. The monoglycosyl glycosphingolipid, glucosyl ceramide, which is the common skeleton of many complex glycosphingolipids including those studied here, was also deuterated at fatty acid C-2 for comparative study in the same matrices. Correlation with spectra of the complex glycolipids demonstrated that, for a given temperature and membrane composition, ceramide backbone conformation was very similar amongst the species studied. Spectral features of GM1 deuterated on terminal NANA and assembled at a membrane surface, were found to be highly consistent with the oligosaccharide conformation determined in studies of GM1 in solution. In contrast, globoside deuterated in the terminal GalNAc residue gave spectra very different from those predicted on the basis of the conformation considered to exist in solution. It seems likely that this result reflects a combination of greater oligosaccharide chain flexibility relative to GM1, and the presence of the membrane environment. Interestingly, although there was highly significant spatial geometry associated with the complex oligosaccharide chains, and although temperature and the presence of cholesterol exert measurable effects on the membrane-inserted portion, these factors had very little impact on the measured spectral parameters associated with the NANA residue of GM1 or the terminal GalNAc residue of globoside. This seems to indicate lack of sensitivity of the complex oligosaccharide chains to conformation and internal motions of the hydrophobic chain segments in these fluid and semi-fluid membranes; and has important implications for mechanisms of crypticity.
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