Journal article
Quantitative mapping of the N-linked sialyloligosaccharides of recombinant erythropoietin: Combination of direct high-performance anion-exchange chromatography and 2-aminopyridine derivatization
Analytical biochemistry, Vol.206(2), pp.278-287
1992
DOI: 10.1016/0003-2697(92)90367-G
PMID: 1443598
Abstract
A rapid quantitative analysis of the sialylated N-linked oligosaccharides of recombinant erythropoietin (EPO) expressed in Chinese hamster ovary (CHO) cells has been developed. The procedure utilizes a glycoamidase (glycopeptidase F) to release all of the N-linked oligosaccharides from the native glycoprotein, followed by direct chromatographic analysis using high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection. The eight sialyloligosaccharides isolated from HPAEC were characterized by derivatizing with 2-aminopyridine followed by two-dimensional HPLC mapping of the pyridylaminated asialooligosaccharides (Tomiya
et al., 1988,
Anal. Biochem. 171, 73–90). Seven kinds of complex-type asialooligosaccharides were identified ranging from a biantennary structure to N-acetyllactosamine-extended tetraantennary structure. Approximately 3% of the terminal galactose residues of the oligosaccharides released from EPO were not sialylated whereas 97% contained an α(2→3)-linked sialic acid. Quantitative oligosaccharide mapping of four different lots of EPO from CHO cells was performed to quantify the molar balance and distribution of the N-linked oligosaccharides. The sialyloligosaccharides were distributed with approximately 5% disialylated (single type), 20% trisialylated (six types), and 75% tetrasialylated (four types) oligosaccharides with an average molar recovery of 85% starting from 750 pmol of EPO.
Details
- Title: Subtitle
- Quantitative mapping of the N-linked sialyloligosaccharides of recombinant erythropoietin: Combination of direct high-performance anion-exchange chromatography and 2-aminopyridine derivatization
- Creators
- Kevin G Rice - Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218 USANoriko Takahashi - Nagoya City University College of Nursing, Nagoya 467, JapanYoshihiro Namiki - Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218 USAAn D Tran - Nagoya City University College of Nursing, Nagoya 467, JapanPeter J Lisi - R. W. Johnson Pharmaceutical Research Institute, Raritan, New Jersey, USAYuan C Lee - Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218 USA
- Resource Type
- Journal article
- Publication Details
- Analytical biochemistry, Vol.206(2), pp.278-287
- DOI
- 10.1016/0003-2697(92)90367-G
- PMID
- 1443598
- NLM abbreviation
- Anal Biochem
- ISSN
- 0003-2697
- eISSN
- 1096-0309
- Publisher
- Elsevier Inc
- Language
- English
- Date published
- 1992
- Academic Unit
- Pharmaceutical Sciences and Experimental Therapeutics; Craniofacial Anomalies Research Center; Medicinal and Natural Products Chemistry
- Record Identifier
- 9984065698402771
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