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Characterization of N-linked oligosaccharides bearing sialyl lewis x moieties on an alternatively glycosylated form of soluble complement receptor type 1 (sCR1).
Biotechnol Appl Biochem. 2000 02; 31(1):5-13.BA

Abstract

We sought to produce a complement inhibitory protein possessing oligosaccharides specifically modified to contain the sialyl Lewis x (sLe(x)) moiety. This modified glycoprotein could combine anti-complement activity with the ability to inhibit selectin-mediated interactions and concentrate this activity to sites of activated endothelium where selectins are upregulated. Soluble complement receptor type 1 (sCR1), previously shown to be effective in inhibiting the complement cascade, was produced in a cell line capable of adding fucose to N-linked oligosaccharides in the alpha1-3 linkage, which is necessary for sLe(x) glycosylation. The glycoprotein purified from these cells was designated sCR1sLe(x), and may prove to be more effective than sCR1 in some clinical applications. Detailed analysis and characterization of sCR1sLe(x) was performed to confirm that the N-linked oligosaccharides possessed sLe(x) moieties and also to determine the extent of sLe(x) glycosylation. The glycoproteins were characterized by oligosaccharide profiling, sequencing, linkage analysis and quantified by differential enzymic digestion, using fluorophore-assisted carbohydrate electrophoresis. The major glycans were identified as biantennary oligosaccharides (including sialylated and non-core fucosylated glycans). The linkages of sialic acid and the branched fucose were analysed by digestion with linkage-specific enzymes and subsequent separation by electrophoresis. All data were consistent with the presence of sLe(x) moieties on the N-linked oligosaccharides of sCR1sLe(x). sCR1sLe(x) is a prime example of a recombinant protein expressed with oligosaccharides engineered for a specific biological function, and produced using a commercially viable method.

Authors+Show Affiliations

AVANT Immunotherapeutics, Inc., 119 Fourth Avenue, Needham, MA 02494, USA.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

10669397

Citation

Picard, M D., et al. "Characterization of N-linked Oligosaccharides Bearing Sialyl Lewis X Moieties On an Alternatively Glycosylated Form of Soluble Complement Receptor Type 1 (sCR1)." Biotechnology and Applied Biochemistry, vol. 31, no. 1, 2000, pp. 5-13.
Picard MD, Pettey CL, Marsh HC, et al. Characterization of N-linked oligosaccharides bearing sialyl lewis x moieties on an alternatively glycosylated form of soluble complement receptor type 1 (sCR1). Biotechnol Appl Biochem. 2000;31(1):5-13.
Picard, M. D., Pettey, C. L., Marsh, H. C., & Thomas, L. J. (2000). Characterization of N-linked oligosaccharides bearing sialyl lewis x moieties on an alternatively glycosylated form of soluble complement receptor type 1 (sCR1). Biotechnology and Applied Biochemistry, 31(1), 5-13.
Picard MD, et al. Characterization of N-linked Oligosaccharides Bearing Sialyl Lewis X Moieties On an Alternatively Glycosylated Form of Soluble Complement Receptor Type 1 (sCR1). Biotechnol Appl Biochem. 2000;31(1):5-13. PubMed PMID: 10669397.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Characterization of N-linked oligosaccharides bearing sialyl lewis x moieties on an alternatively glycosylated form of soluble complement receptor type 1 (sCR1). AU - Picard,M D, AU - Pettey,C L, AU - Marsh,H C, AU - Thomas,L J, PY - 2000/2/11/pubmed PY - 2000/4/15/medline PY - 2000/2/11/entrez SP - 5 EP - 13 JF - Biotechnology and applied biochemistry JO - Biotechnol Appl Biochem VL - 31 IS - 1 N2 - We sought to produce a complement inhibitory protein possessing oligosaccharides specifically modified to contain the sialyl Lewis x (sLe(x)) moiety. This modified glycoprotein could combine anti-complement activity with the ability to inhibit selectin-mediated interactions and concentrate this activity to sites of activated endothelium where selectins are upregulated. Soluble complement receptor type 1 (sCR1), previously shown to be effective in inhibiting the complement cascade, was produced in a cell line capable of adding fucose to N-linked oligosaccharides in the alpha1-3 linkage, which is necessary for sLe(x) glycosylation. The glycoprotein purified from these cells was designated sCR1sLe(x), and may prove to be more effective than sCR1 in some clinical applications. Detailed analysis and characterization of sCR1sLe(x) was performed to confirm that the N-linked oligosaccharides possessed sLe(x) moieties and also to determine the extent of sLe(x) glycosylation. The glycoproteins were characterized by oligosaccharide profiling, sequencing, linkage analysis and quantified by differential enzymic digestion, using fluorophore-assisted carbohydrate electrophoresis. The major glycans were identified as biantennary oligosaccharides (including sialylated and non-core fucosylated glycans). The linkages of sialic acid and the branched fucose were analysed by digestion with linkage-specific enzymes and subsequent separation by electrophoresis. All data were consistent with the presence of sLe(x) moieties on the N-linked oligosaccharides of sCR1sLe(x). sCR1sLe(x) is a prime example of a recombinant protein expressed with oligosaccharides engineered for a specific biological function, and produced using a commercially viable method. SN - 0885-4513 UR - https://www.unboundmedicine.com/medline/citation/10669397/Characterization_of_N_linked_oligosaccharides_bearing_sialyl_lewis_x_moieties_on_an_alternatively_glycosylated_form_of_soluble_complement_receptor_type_1__sCR1__ L2 - https://onlinelibrary.wiley.com/resolve/openurl?genre=article&sid=nlm:pubmed&issn=0885-4513&date=2000&volume=31&issue=1&spage=5 DB - PRIME DP - Unbound Medicine ER -