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Determination of PEGylation homogeneity of polyethylene glycol-modified canine uricase.
- Source :
-
Electrophoresis [Electrophoresis] 2021 Mar; Vol. 42 (6), pp. 693-699. Date of Electronic Publication: 2020 Dec 16. - Publication Year :
- 2021
-
Abstract
- Polyethylene glycol-modified canine uricase (PEG-UHC) was prepared by modifying the ε-amino group of lysine residues on the canine uricase (UHC) protein to near-saturation with 5 kDa monomethoxyl-polyethylene glycol succinimide (mPEG-SPA-5k). In order to accurately determine the PEGylation uniformity of PEG-UHC, CZE, 3-8% gradient gel SDS-PAGE, and imaging CIEF (iCIEF) analyses were compared. CZE could not effectively separate PEG-UHC proteins with different degrees of modification, 3-8% gradient gel SDS-PAGE could separate PEG-UHC into seven gel bands; however, most of the gel bands were smeared or blurred, and the separation of PEG-UHC samples by iCIEF was significantly better than that by 3-8% gradient gel SDS-PAGE. Under denatured conditions, iCIEF separated 12 pI peaks, and could also accurately quantify the relative monomer PEG-UHC content. More than 85% of the total monomeric PEG-UHC was conjugated with 7-12 PEG molecules; of this 85%, approximately 40% was conjugated with 9-10 PEG molecules. These results demonstrated that iCIEF exhibits good potential for determining the PEGylation homogeneity of PEGylated protein drugs.<br /> (© 2020 Wiley-VCH GmbH.)
Details
- Language :
- English
- ISSN :
- 1522-2683
- Volume :
- 42
- Issue :
- 6
- Database :
- MEDLINE
- Journal :
- Electrophoresis
- Publication Type :
- Academic Journal
- Accession number :
- 33247595
- Full Text :
- https://doi.org/10.1002/elps.202000268