Binding of Ca2+, Mg2+, and heparin by human serum amyloid P component in affinity capillary electrophoresis

Human serum amyloid P component (SAP) is a glycoprotein circulating in the blood and found in association with all types of amyloid (malfolded potein aggregates) examined so far. Despite uncertainties regarding the precise function of SAP in vivo, the lectin-like properties of this Ca2+-activated protein with affinity for anionic saccharides and malfolded proteins are well known. The propensity to form homomeric penta- or decamers in solution and the selfaggregation in the presence of Ca2+ as well as the tendency of SAP to attach to uncoated fused silica have precluded the analysis of SAP by microelectrophoretic methods. We now work out conditions to characterize the binding of Ca2+ and Mg2+ and the binding of heparin to SAP in the presence of divalent metal ions by ACE. The results show a strong binding of heparin (sub-mu M apparent dissociation constants) even in the abscence of Ca2+ at low ionic strength, pH 8.2. Also, a selective interaction with Ca2+ compared with Mg2+ is demonstrated. The approach will further the use of microelectrophoretic methods to examine the interactions of SAP with ligands of putative pathophysiological relevance such as lipopolysaccharides and misfolded proteins.