The conformational effects of N-glycosylation on the tailpiece from serum IgM.
1H-NMR spectroscopy has been used to study the conformation and dynamics of the isolated tailpiece from human serum immunoglobulin M, a 22-residue peptide containing a single asparagine glycosylation site. The peptide is isolated as a set of glycoforms, varying only in the sequence of the oligosaccharide attached at the glycosylation site. The oligosaccharides present have the general formula (Man)n(GlcNAc)2, with 45% having n = 6, 45% having n = 8 and 10% having n = 7 and/or 9. They have been identified and their NMR parameters compared to those found for the isolated oligosaccharides in free solution. The conformation and dynamics of the peptide component have also been studied, using NOE data and hydrogen-exchange experiments, and the results compared to those obtained from the aglycosyl peptide of the same sequence. The presence of the peptide is found to have no measurable effect on the conformation of the oligosaccharides. However, the presence of oligosaccharide causes a decrease in the conformational mobility of the backbone and sidechains of the peptide in the region of the glycosylation site. This is proposed to result from interactions between the oligosaccharide core and the amino acid side chains. Further, the conformation of the N-glycosidic linkage has been shown to be both rigid and planar. Thus, the conformational space available to an N-linked oligosaccharide in a glycoprotein relative to the protein may depend to a large extent upon the flexibility of the asparagine side chain. Various roles for the different glycoforms of the tail peptide are discussed.
Wormald
M R
MR
Department of Biochemistry, University of Oxford, England.
Wooten
E W
EW
Bazzo
R
R
Edge
C J
CJ
Feinstein
A
A
Rademacher
T W
TW
Dwek
R A
RA
D000595:Amino Acid Sequence; D002236:Carbohydrate Conformation; D006031:Glycosylation; D006801:Humans; D007075:Immunoglobulin M / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry; D011487:Protein Conformation
D007075:Immunoglobulin M; D009844:Oligosaccharides