Analysis of the site-specific asparagine-linked glycosylation of recombinant human coagulation factor VLLa by glycosidase digestions, liquid chromatography, and mass spectrometry.
The two asparagine-linked glycosylation sites of recombinant coagulation factor VIIa have been characterized by glycosidase digestions, size-exclusion chromatography (SEC), and mass spectrometry (MS). Nine structures were characterized as core fucosylated bi- and triantennary structures with 0-3 sialic-acid residues, which were alpha2-3 linked to galactose exclusively. Three of the structures had one or two galactose residues substituted by N-acetylgalactosamine. Significant differences were found between the oligosac-charide profiles for the two glycosylation sites in rFVIIa. At Asn322, the degree of sialylation was lower and higher amounts of structures containing N-acetylgalactosamine were found compared to Asn l45.
Kristian Klausen
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Department of Protein Chemistry Ge, Novo Nordisk A/S, Niels Steensensvej 1, 2820, Gentofte, Denmark, nkk@novo.dk.
Bayne
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Palm
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