Kr. Barber et al., BEHAVIOR OF COMPLEX OLIGOSACCHARIDES AT A BILAYER-MEMBRANE SURFACE - PROBED BY H-2-NMR, Biochimica et biophysica acta. Biomembranes, 1190(2), 1994, pp. 376-384
Deuterium wideline NMR was used in an attempt to directly assess oligo
saccharide arrangement and motional characteristics of complex glycosp
hingolipids dispersed as minor components in phospholipid membranes. A
convenient, general synthetic approach was developed which involved r
eplacement of the acetate group of amido sugars with deuteroacetate (-
COCD3). This provided excellent signal-to-noise when applied to the te
rminal GalNAc residue of globoside, and the terminal NANA residue of G
M1. Simultaneously, globoside and GM1 fatty acids were replaced with s
tearic acid deuterated at C-2 - a probe location sensitive to glycolip
id hydrophobic backbone orientation and rigid body motion. Deuterated
GM1 and globoside were studied by H-2-NMR in bilayers of 1-palmitoyl-2
-oleoyl phosphatidylcholine, in the presence and absence of physiologi
cal quantities of cholesterol. The monoglycosyl glycosphingolipid, glu
cosyl ceramide, which is the common skeleton of many complex glycosphi
n-golipids 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 temp
erature and membrane composition, ceramide backbone conformation was v
ery similar amongst the species studied. Spectral features of GM1 deut
erated on terminal NANA and assembled at a membrane surface, were foun
d to be highly consistent with the oligosaccharide conformation determ
ined 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 solu
tion. It seems likely that this result reflects a combination of great
er 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 oligosacchar
ide chains, and although temperature and the presence of cholesterol e
xert measurable effects on the membrane-inserted portion, these factor
s had very little impact on the measured spectral parameters associate
d with the NANA residue of GM1 or the terminal GalNAc residue of globo
side. This seems to indicate lack of sensitivity of the complex oligos
accharide chains to conformation and internal motions of the hydrophob
ic chain segments in these fluid and semi-fluid membranes; and has imp
ortant implications for mechanisms of crypticity.