Chemical crosslinking and mass spectrometric identification of interaction sites within soluble aggregate of protein therapeutics.
Protein aggregation presents a major challenge to bioengineering. In the present study, chemical crosslinking and mass spectrometry are employed to probe the interaction amongst soluble oligomer formed by Fc molecule of monoclonal antibody. Aggregation was induced by thermal stress at 42°C for 6h under physiological pH. Contacting residues on adjacent molecules were captured with bifunctional crosslinker BS3, followed by mass spectrometric identification of the crosslinked sites. The approach provides site-specific information regarding the binding interface that would have broad application in the field.
Massoverz Technologies, Albany, NY 12203, USA.
Alan
A
Zhao
Gang
G
Hao
Jane
J
Gu
D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D002850:Chromatography, Gel; D006224:Cricetinae; D003412:Cricetulus; D003432:Cross-Linking Reagents / Q000737:chemistry*; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry* / Q000235:genetics; D007155:Immunologic Factors / Q000737:chemistry* / Q000235:genetics; D013058:Mass Spectrometry; D010449:Peptide Mapping / Q000379:methods*; D055503:Protein Multimerization; D012995:Solubility
D003432:Cross-Linking Reagents; D007141:Immunoglobulin Fc Fragments; D007155:Immunologic Factors