HYDROGEN/DEUTERIUM EXCHANGE REVEALS THE CONFORMATIONAL CHANGES OF HUMAN a1-ACID GLYCOPROTEIN UPON GLYCOSYLATION
Richard Y.-C. Huang and Jeffrey W. Hudgens
Human a1-Acid Glycoprotein (AGP) or orosomucoid, a 40 kDa acute phase glycoprotein, presents predominantly in blood. The glycosylation and serum concentration of AGP change in response to tissue injury, inflammation or infection, and these changes are highly correlated with hepatic synthesis. With its ability to bind basic, lipophilic, and acidic drugs, AGP has been served as a suitable drug carrier. It has been shown that the drug-binding activities of AGP depend strongly on its carbohydrate composition. The details of the effects of glycosylation on the AGP conformation, however, are unknown. Here we report the use of hydrogen/deuterium exchange (H/DX) coupled with mass spectrometry to elucidate the changes in conformation and dynamics of AGP upon different types and degree of glycosylation. Our peptide-level H/DX not only reveals the dynamics of AGP variants, but also demonstrates that glycosylation modulates dynamics of five peptide regions (residue 9-18, 44-48, 50-61, 80-89, and 114-127) surrounding the ligand-binding cavity of AGP, which must cause significant impact on its drug-binding activities and immunomodulatory.