A strategy for characterization of polyethylene glycol-derivatized proteins. A mass spectrometric analysis of the attachment sites in polyethylene glycol-derivatized superoxide dismutase
Vestling, M.M.; Murphy, C.M.; Keller, D.A.; Fenselau, C.; Dedinas, J.; Ladd, D.L.; Olsen, M.A.
Drug Metabolism and Disposition the Biological Fate of Chemicals 21(5): 911-917
1993
ISSN/ISBN: 0090-9556 PMID: 7902255 Document Number: 416225
Base treatment of polyethylene glycol-derivatized superoxide dismutase in which the polyethylene glycol is linked to the protein via a succinyl bridge, removes the polyethylene glycol leaving a succinyl marker. Exhaustive succinylation with d-4-succinic anhydride completes the derivatization in order to minimize fractionation in proteolysis, chromatography, and desorption in the mass spectrometer. Production of peptides from the derivatized protein for high resolution and high-resolution tandem MS allows identification of the site that had been derivatized by polyethylene glycol and the determination of the amount of polyethylene glycol originally at each site. The mass spectrometric strategy outlined herein can be applied to other proteins derivatized for therapeutic administration.