Effect of genetic background on glycosylation heterogeneity in human antithrombin produced in the mammary gland of transgenic goats.
Glycosylation is involved in the correct folding, targeting, bioactivity and clearance of therapeutic glycoproteins. With the development of transgenic animals as expression systems it is important to understand the impact of different genetic backgrounds and lactations on glycosylation. We have evaluated the glycosylation of recombinant antithrombin produced in several transgenic goat lines, from cloned animals and from different types of lactation including induced lactations. Our results show glycosylation patterns from the protein expressed in animals, derived from the same founder goat, are mostly comparable. Furthermore, the protein expressed in two cloned goats had highly consistent oligosaccharide profiles and similar carbohydrate composition. However, there were significantly different oligosaccharide profiles from the proteins derived from different founder goats. Artificial induction of lactation did not have significant effects on overall carbohydrate structures when compared to natural lactation. The only major difference was that recombinant antithrombin from induced lactations contained a slightly higher ratio of N-acetylneuraminic acid to N-glycolylneuraminic acid and less amount of oligosaccharides containing N-glycolylneuraminic acid. The oligosaccharides from all animals were a mixture of high mannose-, hybrid- and complex-type oligosaccharides. Sialic acid was present as alpha-2,6-linkage and no alpha-1,3-linked galactose was observed. These results indicate that transgenic animals with closely related genetic backgrounds express recombinant protein with comparable glycosylation.
Zhou
Qun
Q
Therapeutic Protein Research, Genzyme Corporation, P.O. Box 9322, Framingham, MA 01701-9322, USA. qun.zhou@genzyme.com
Kyazike
Josephine
J
Echelard
Yann
Y
Meade
Harry M
HM
Higgins
Elizabeth
E
Cole
Edward S
ES
Edmunds
Tim
T
D000818:Animals; D030801:Animals, Genetically Modified; D000991:Antithrombins / Q000096:biosynthesis* / Q000737:chemistry; D005260:Female; D006031:Glycosylation; D006041:Goats / Q000235:genetics*; D006801:Humans; D007774:Lactation; D008321:Mammary Glands, Animal / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry; D011994:Recombinant Proteins / Q000096:biosynthesis*
D000991:Antithrombins; D009844:Oligosaccharides; D011994:Recombinant Proteins