Structural study of the sugar moieties of monoclonal antibodies secreted by human-mouse hybridoma.
Six monoclonal antibodies, three each of human IgG1 and IgG2 subclasses, were obtained from human-mouse hybridomas. Structural study of their asparagine-linked sugar chains was performed to elucidate the regulatory mechanism of secreted monoclonal IgG glycosylation. The sugar moieties were quantitatively released as oligosaccharides from the polypeptide backbone by hydrazinolysis. They were converted into radioactive oligosaccharides by NaB3H4 reduction after N-acetylation. Structural study of each oligosaccharide by lectin affinity column chromatography, sequential exoglycosidase digestion, and methylation analysis indicated that almost all of them were biantennary complex-type sugar chains containing Man alpha 1----6(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4 (+/- Fuc alpha 1----6)GlcNAc as core structures. Bisecting N-acetylglucosamine residue, which is present in human IgG but not in mouse IgG, could not be detected at all. The molar ratio of each oligosaccharide from the six IgG samples was different. However, no subclass specificity was detected except that all IgG1 contained neutral, mono-, and disialylated sugar chains, whereas IgG2 did not contain disialylated ones. The molar ratio of N-acetylneuraminic acid to N-glycolylneuraminic acid was also different for each IgG. All six IgGs contained monoantennary complex-type and high mannose-type oligosaccharides which had never been detected in serum IgGs of various mammals so far investigated. These results indicated that the processing of asparagine-linked sugar chains of IgG is less complete in human-mouse hybridoma than in human or mouse B cells, and that the glycosylation machinery of the mouse cells is dominant in the hybrid cells.
Tandai
M
M
Department of Biochemistry, University of Tokyo, Japan.
Endo
T
T
Sasaki
S
S
Masuho
Y
Y
Kochibe
N
N
Kobata
A
A
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002240:Carbohydrate Sequence; D005591:Chemical Fractionation; D002845:Chromatography / Q000379:methods; D004590:Electrophoresis, Paper; D006801:Humans; D006825:Hybridomas / Q000378:metabolism*; D007074:Immunoglobulin G / Q000737:chemistry; D051379:Mice; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D009844:Oligosaccharides