title: N-Glycosylation profiling of recombinant mouse extracellular superoxide dismutase produced in Chinese hamster ovary cells.
abstract: Extracellular superoxide dismutase (EC-SOD), the major SOD isoenzyme in biological fluids, is known to be N-glycosylated and heterogeneous as was detected in most glycoproteins. However, only one N-glycan structure has been reported in recombinant human EC-SOD produced in Chinese hamster ovary (CHO) cells. Thus, a precise N-glycan profile of the recombinant EC-SOD is not available. In this study, we report profiling of the N-glycan in the recombinant mouse EC-SOD produced in CHO cells using high-resolution techniques, including the liberation of N-glycans by treatment with PNGase F, fluorescence labeling by pyridylamination, characterization by anion-exchange, normal and reversed phase-HPLC separation, and mass spectrometry. We succeeded in identifying 26 different types of N-glycans in the recombinant enzyme. The EC-SOD N-glycans were basically core-fucosylated (98.3% of the total N-glycan content), and were high mannose sugar chain, and mono-, bi-, tri-, and tetra-antennary complex sugar chains exhibiting varying degrees of sialylation. Four of the identified N-glycans were uniquely modified with a sulfate group, a Lewis(x) structure, or an α-Gal epitope. The findings will shed new light on the structure-function relationships of EC-SOD N-glycans.
authors.affiliation: Department of Disease Glycomics (Seikagaku Corporation), The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka, 567-0047, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Korekane
authors.forename: Atsuko
authors.initials: A
authors.lastname: Korekane
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: Masaki
authors.initials: M
authors.lastname: Kato
authors.forename: Yasuhide
authors.initials: Y
authors.lastname: Miyamoto
authors.forename: Akio
authors.initials: A
authors.lastname: Matsumoto
authors.forename: Tomoko
authors.initials: T
authors.lastname: Hasegawa
authors.forename: Keiichiro
authors.initials: K
authors.lastname: Suzuki
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Taniguchi
authors.forename: Tomomi
authors.initials: T
authors.lastname: Ookawara
mesh_terms: D000818:Animals; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D003412:Cricetulus; D006031:Glycosylation; D013058:Mass Spectrometry; D051379:Mice; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D013482:Superoxide Dismutase