1. Characterization of a monoPEG20000-Endostar
- Author
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Tong, Yue, Zhong, Kai, Tian, Hong, Gao, Xiangdong, Xu, Xiangyang, Yin, Xiaojin, and Yao, Wenbing
- Subjects
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LUNG cancer , *POLYETHYLENE glycol , *ALDEHYDES , *ION exchange chromatography , *BARIUM compounds , *WESTERN immunoblotting , *THERAPEUTICS - Abstract
Abstract: In this study, we investigated the PEG attachment site of mono-PEGylated Endostar, a modified recombinant human endostatin approved in China for lung cancer. N-terminal site-directed mono-PEGylation of Endostar was accomplished using mPEG-propionaldehyde derivatives (Mw=20kDa) under slightly acidic pH conditions (pH 5.5). One-step cation exchange chromatography was used to purify the mono-PEGylated Endostar. Following tryptic digestion, the peptide fragment containing PEG was separated by SDS-PAGE. Barium iodide staining and Western blotting were used to detect the PEG moiety and the N-terminus of Endostar, respectively. The peptide fragment stained by barium iodide showed a positive response to anti-(His) 6 mAb, demonstrating that PEG was located at the N-terminus of Endostar. LC/MS was applied to verify the occurrence of mono-PEGylation at the N-terminus of Endostar. [Copyright &y& Elsevier]
- Published
- 2010
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