Stabilization of bacterially expressed erythropoietin by single site-specific introduction of short branched PEG chains at naturally occurring glycosylation sites
| Autor: | Christian P. R. Hackenberger, Thomas Brunner, Nicole Nischan, Marina Rubini, Carina Seitz, Sergej Schwagerus, Eugenia Hoffmann, Katharina Streichert |
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| Rok vydání: | 2016 |
| Předmět: |
Glycosylation
Protein Conformation Gene Expression 02 engineering and technology Polyethylene glycol 010402 general chemistry 01 natural sciences Cell Line Polyethylene Glycols chemistry.chemical_compound Protein structure PEG ratio Escherichia coli medicine Animals Humans Amino Acid Sequence Amino Acids Erythropoietin Molecular Biology Cell Proliferation chemistry.chemical_classification Bioconjugation Molecular Structure Protein Stability 021001 nanoscience & nanotechnology Recombinant Proteins 0104 chemical sciences Amino acid chemistry Biochemistry ddc:540 PEGylation 0210 nano-technology Biotechnology medicine.drug |
| Zdroj: | Molecular BioSystems, 12: 1750-1755 |
| ISSN: | 1742-2051 1742-206X |
| DOI: | 10.1039/c5mb00857c |
| Popis: | The covalent attachment of polyethylene glycol (PEG) to therapeutic proteins can improve their physicochemical properties. In this work we utilized the non-natural amino acid p-azidophenylalanine (pAzF) in combination with the chemoselective Staudinger-phosphite reaction to install branched PEG chains to recombinant unglycosylated erythropoietin (EPO) at each single naturally occurring glycosylation site. PEGylation with two short 750 or 2000 Da PEG units at positions 24, 38, or 83 significantly decreased unspecific aggregation and proteolytic degradation while biological activity in vitro was preserved or even increased in comparison to full-glycosylated EPO. This site-specific bioconjugation approach permits to analyse the impact of PEGylation at single positions. These results represent an important step towards the engineering of site-specifically modified EPO variants from bacterial expression with increased therapeutic efficacy. published |
| Databáze: | OpenAIRE |
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