Structure of the oligosaccharide of hen phosvitin as determined by two-dimensional 1H NMR of the intact glycoprotein.
The major form of the oligosaccharide of hen phosvitin was studied with two-dimensional 1H NMR of the intact glycoprotein. Its structure was determined from an analysis of the chemical shifts of the structural reporter groups, and it was further confirmed by comparison to several related model oligosaccharides. The oligosaccharide is N-linked and is present in a 1:1 stoichiometry to the protein. It has a complex type 1 triantennary structure with two NeuAc alpha 2,6Gal beta 1,4GlcNAc beta 1,2 arms linked to the Man-4 and Man-4' and a third Gal beta 1, 4GlcNAc beta 1,4 arm attached to the Man-4. The oligosaccharide contains the common core sequence which is present in all N-linked glycoproteins [Man alpha 1,3(Man alpha 1,6)-Man beta 1,4GlcNAc beta 1,4GlcNAc beta 1,N]. In the course of this study, we have found that unique spin systems for the GlcNAc and NeuAc are obtained for spectra recorded in 90% H2O. Their NH peaks were assigned at low pH, and these assignments proved useful for confirming the identity of cross-peaks in the anomeric region. In addition, the protons of GlcNAc-1 could be correlated to the NH of the asparagine link. The cross-peak patterns determined in phase-sensitive 2D experiments for the H1,H2 protons have a different appearance for each type of monosaccharide, and this information was also used for making first-order assignments. A comparison with model compounds suggests that the solution conformation of the oligosaccharide is not affected by its attachment to the protein.
Brockbank
R L
RL
Department of Biological Sciences, University of Calgary, Alberta, Canada.
Vogel
H J
HJ
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002645:Chickens; D004527:Egg Proteins; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009844:Oligosaccharides; D010774:Phosvitin
D004527:Egg Proteins; D009844:Oligosaccharides; D010774:Phosvitin