Document Detail


Metabolic control of recombinant protein N-glycan processing in NS0 and CHO cells.
MedLine Citation:
PMID:  11257601     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Chinese hamster ovary and murine myeloma NS0 cells are currently favored host cell types for the production of therapeutic recombinant proteins. In this study, we compared N-glycan processing in GS-NS0 and GS-CHO cells producing the same model recombinant glycoprotein, tissue inhibitor of metalloproteinases 1. By manipulation of intracellular nucleotide-sugar content, we examined the feasibility of implementing metabolic control strategies aimed at reducing the occurrence of murine-specific glycan motifs on NS0-derived recombinant proteins, such as Galalpha1,3Galbeta1,4GlcNAc. Although both CHO and NS0-derived oligosaccharides were predominantly of the standard complex type with variable sialylation, 30% of N-glycan antennae associated with NS0-derived TIMP-1 terminated in alpha1,3-linked galactose residues. Furthermore, NS0 cells conferred a greater proportion of terminal N-glycolylneuraminic (sialic) acid residues as compared with the N-acetylneuraminic acid variant. Inclusion of the nucleotide-sugar precursors, glucosamine (10 mM, plus 2 mM uridine) and N-acetylmannosamine (20 mM), in culture media were shown to significantly increase the intracellular pools of UDP-N-acetylhexosamine and CMP-sialic acid, respectively, in both NS0 and CHO cells. The elevated UDP-N-acetylhexosamine content induced by the glucosamine/uridine treatment was associated with an increase in the antennarity of N-glycans associated with TIMP-1 produced in CHO cells but not N-glycans associated with TIMP-1 from NS0 cells. In addition, elevated UDP-N-acetylhexosamine content was associated with a slight decrease in sialylation in both cell lines. The elevated CMP-sialic acid content induced by N-acetylmannosamine had no effect on the overall level of sialylation of TIMP-1 produced by both CHO and NS0 cells, although the ratio of N-glycolylneuraminic acid:N-acetylneuraminic acid associated with NS0-derived TIMP-1 changed from 1:1 to 1:2. These data suggest that manipulation of nucleotide-sugar metabolism can promote changes in N-glycan processing that are either conserved between NS0 and CHO cells or specific to either NS0 cells or CHO cells.
Authors:
K N Baker; M H Rendall; A E Hills; M Hoare; R B Freedman; D C James
Publication Detail:
Type:  Comparative Study; Journal Article    
Journal Detail:
Title:  Biotechnology and bioengineering     Volume:  73     ISSN:  0006-3592     ISO Abbreviation:  Biotechnol. Bioeng.     Publication Date:  2001 May 
Date Detail:
Created Date:  2001-03-21     Completed Date:  2001-06-14     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  7502021     Medline TA:  Biotechnol Bioeng     Country:  United States    
Other Details:
Languages:  eng     Pagination:  188-202     Citation Subset:  IM    
Copyright Information:
Copyright 2001 John Wiley & Sons, Inc.
Affiliation:
Department of Biosciences, University of Kent, Canterbury, Kent CT2 7NJ, UK.
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MeSH Terms
Descriptor/Qualifier:
Animals
Bioreactors
CHO Cells
Cell Division / drug effects
Chromatography, High Pressure Liquid
Cricetinae
Galactose / metabolism
Glucosamine / pharmacology
Glycosylation / drug effects
Hexosamines / pharmacology
Mice
N-Acetylneuraminic Acid / metabolism
Nucleotides / metabolism
Polysaccharides / chemistry,  metabolism*
Recombinant Proteins / metabolism
Sialyltransferases / metabolism
Tissue Inhibitor of Metalloproteinase-1 / chemistry,  metabolism*
Tumor Cells, Cultured
Chemical
Reg. No./Substance:
0/Hexosamines; 0/Nucleotides; 0/Polysaccharides; 0/Recombinant Proteins; 0/Tissue Inhibitor of Metalloproteinase-1; 131-48-6/N-Acetylneuraminic Acid; 26566-61-0/Galactose; 3416-24-8/Glucosamine; 4773-29-9/N-acetylmannosamine; EC 2.4.99.-/Sialyltransferases

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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