ISSN:
1573-4986
Keywords:
biotechnological antibody
;
chimeric IgG1
;
N-glycosylation
;
cell subclones
;
glycoprotein
Source:
Springer Online Journal Archives 1860-2000
Topics:
Chemistry and Pharmacology
Notes:
Abstract Two chimeric human/murine monoclonal antibodies were constructed by substitution of the murine constant regions with human γ1 and κ constant regions for heavy and light chains, respectively. The chimeric human/murine molecules are anti-idiotypic antibodies, meaning that they were directed against the antigen binding site in the variable region of another antibody. Antibody batches were produced under identical production conditions, using two selected SP2/0 myeloma cell subclones, which produce chimeric antibodies with different variable regions, but identical constant regions. Several samples were collected during the production of the antibodies in hollow-fibre reactors. The heavy chain, but not the light chain, of the two different chimeric IgG1 antibodies is glycosylated. Structural analysis of the enzymically released N-linked carbohydrate chains by1H-NMR spectroscopy, as well as by chromatographic profiling, demonstrated that the collection of N-glycans comprises a small amount of monoantennary, and for the greater part diantennary structures. The N-glycans are completely (α1 →6)-flucosylated at the innermost GlcNAc residue. The antennae of the neutral diantennary N-glycans are built up from GlcNAcβ1→2, Galβ1 → 4GlcNAcβ1 → 2 or Galα1 → 3Gα1β1 → 4GlcNAcβ1 → 2 elements, whereas the antennae of the neutral monoantennary carbohydrate chains have only (β1 → 2)-linked GlcNAc residues. Galactosylation of the GlcNAcβ1 → 2Manα1 → 6 branch occurs four times more frequently than that of the GlcNAcβ1 → 2Manα1 → 3 branch, independently of the production batch. A small amount of the diantennary N-glycans are mono- or disialylated, carryingN-acetylneuraminic acid (Neu5Ac) orN-glycolylneuraminic acid (Neu5Gc), exclusively (α2 → 6)-linked to βGal. Analysis of the different production batches demonstrates that the structures of the N-linked carbohydrate chains are identical in the two chimeric antibodies, but that the relative amounts of the major oligosaccharide components, the degree of sialylation and the molar ratio of Neu5Ac to Neu5Gc varies with the SP2/0 cell subclone, and only slightly with cell age.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00731335
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