Characterization of a monoPEG20000-Endostar.

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=20 kDa) 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.

• Authors: Yue Tong, Kai Zhong, Hong Tian, Xiangdong Gao, Xiangyang Xu, Xiaojin Yin, Wenbing Yao
• Journal: International journal of biological macromolecules (Int J Biol Macromol) Vol. 46 Issue 3 Pg. 331-6 (Apr 01 2010) ISSN: 1879-0003 [Electronic] Netherlands
• PMID: 20122957 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright 2010 Elsevier B.V. All rights reserved.

Chemical References

• Endostatins
• Peptides
• Recombinant Proteins
• Polyethylene Glycols
• Trypsin
• endostar protein
• mono-PEGylated Endostar

Topics

• Amino Acid Sequence
• Blotting, Western
• Chromatography, High Pressure Liquid
• Chromatography, Liquid
• Chromatography, Reverse-Phase
• Electrophoresis, Polyacrylamide Gel
• Endostatins (analysis, chemistry, isolation & purification)
• Humans
• Molecular Sequence Data
• Peptides (chemistry)
• Polyethylene Glycols (analysis, chemistry, isolation & purification)
• Recombinant Proteins
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
• Trypsin (metabolism)