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RCA-I-resistant CHO mutant cells have dysfunctional GnT I and expression of normal GnT I in these mutants enhances sialylation of recombinant erythropoietin

Paper ID Volume ID Publish Year Pages File Format Full-Text
31726 44833 2010 9 PDF Available
Title
RCA-I-resistant CHO mutant cells have dysfunctional GnT I and expression of normal GnT I in these mutants enhances sialylation of recombinant erythropoietin
Abstract

A large number of CHO glycosylation mutants were isolated by Ricinus communis agglutinin-I (RCA-I). Complementation tests revealed that all these mutant lines possessed a dysfunctional N-acetylglucosaminyltransferase I (GnT I) gene. Sequencing analyses on the GnT I cDNAs isolated from 16 mutant lines led to the identification of nine different single base pair mutations. Some mutations result in a premature stop codon whereas others cause a single amino acid substitution in the GnT I protein. Interestingly, expression of the normal GnT I cDNA in mutant cells resulted in enhanced sialylation of N-glycans. The sialylation of recombinant erythropoietin (EPO) produced in mutant cells that were co-transfected with GnT I was enhanced compared to that of EPO produced in wild type CHO cells. The enhanced sialylation of EPO produced by JW152 cells in the presence of GnT I over CHO-K1 cells is a result of increased sialylated glycan structures with higher antennary branching. These findings represent a new strategy that may be utilized by the biotechnology industry to produce highly sialylated therapeutic glycoproteins.

Keywords
CHO glycosylation mutants; Sialylation; Ricinus communis agglutinin-I (RCA-I); N-acetylglucosaminyltransferase I (GnT I); Recombinant erythropoietin (EPO); Isoelectric focusing (IEF)
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RCA-I-resistant CHO mutant cells have dysfunctional GnT I and expression of normal GnT I in these mutants enhances sialylation of recombinant erythropoietin
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Publisher
Database: Elsevier - ScienceDirect
Journal: Metabolic Engineering - Volume 12, Issue 4, July 2010, Pages 360–368
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
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Any Questions? feel free to contact us