A Proteomic Approach for Identifying Cellular Proteins Interacting with Erythropoietin in Recombinant Chinese Hamster Ovary Cells

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Identification of the cellular proteins interacting with incompletely folded and unfolded forms of erythropoietin (EPO) in recombinant CHO (rCHO) cells leads to better insight into the possible genetic manipulation approaches for increasing EPO production. To do so, a pull-down assay was performed with dual-tagged (N-terminal GST- and C-terminal hexahistidine-tagged) EPO expressed in E. coli as bait proteins and cell lysates of rCHO cells (DG44) as prey proteins. Cellular proteins interacting with dual-tagged EPO were then resolved by two-dimensional gel electrophoresis (2DE) and identified by MALDI-TOF MS/MS. A total of 27 protein spots including glucose-regulated protein 78 (GRP78) were successfully identified. Western blot analysis of GRP78 confirmed the results of the MS analyses. Taken together, a pull-down assay followed by a proteomic approach is found to be an efficient means to identify cellular proteins interacting with foreign protein in rCHO cells. (C) 2009 American Institute of Chemical Engineers Biotechnol. Prog., 26: 246-251, 2010
Publisher
AMER CHEMICAL SOC
Issue Date
2010-01
Language
English
Article Type
Article
Keywords

CHO-CELLS; 2-DIMENSIONAL ELECTROPHORESIS; DISULFIDE-ISOMERASE; SODIUM-BUTYRATE; PRODUCTIVITY; IDENTIFICATION; EXPRESSION; ANTIBODY; OVEREXPRESSION; THROMBOPOIETIN

Citation

BIOTECHNOLOGY PROGRESS, v.26, pp.246 - 251

ISSN
8756-7938
DOI
10.1002/btpr.323
URI
http://hdl.handle.net/10203/100973
Appears in Collection
BS-Journal Papers(저널논문)
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