Elucidating the Structural Chemistry of Glycosaminoglycan Recognition by Protein C Inhibitor

Glycosaminoglycans (GAGs) including heparin accelerate the inhibition of serine proteases by serine protease inhibitors (serpins), an essential process in regulating blood coagulation. To analyze the molecular basis for GAG recognition by the plasma serpin protein C inhibitor (PCI; also known as plasminogen activator inhibitor 3), we have constructed a complete, energy-minimized, three-dimensional model of PCI by using the structure of homologous α1-antitrypsin as a template. Sequence analysis, hydrogenbonding environment, and shape complementarity suggested that the N-terminal residues of PCI, which are not homologous to those of α1-antitrypsin, form an amphipathic α-helix, here designated A + since it precedes the α1-antitrypsin A helix. Electrostatic calculations revealed a single, highly positive surface region arising from both the A+ and H helices, suggesting that this two-helix motif is required for GAG binding by PCI. The dominant role of electrostatic interactions in PCI-heparin binding was confirmed by the strong ionic strength dependence of heparin stimulation. The involvement of the A+ helix in heparin binding was verified by demonstrating that an anti-PCI antibody that specifically binds the A+ peptide blocks heparin binding.