Abstract Number: PB1930
Meeting: ISTH 2020 Congress
Background: β2GpI is an abundant plasma protein of yet unknown function and the main autoantigen in the antiphospholipid syndrome (APS). We have recently discovered that β2GpI may function as a mild anticoagulant, as it binds to α-Thrombin (αT) exosite-2, impairing binding to GpIbα platelet receptor [Pozzi et al., JTH (2013); Acquasaliente et al., Biochem J (2016)]. Earlier studies have shown that β2GpI Domain-I is the major epitope for APS autoantibodies.
Aims: Check whether β2GpI interacts with fibrinogen (Fb); identify the binding regions on β2GpI and Fb involved in β2GpI-Fb complex formation; study the effect of β2GpI binding to Fb on fibrin structure.
Methods: Binding measurement were performed by fluorescence and Surface Plasmon Resonance (SPR). Fibrin structure was studied by turbidimetry, Dynamic Light Scattering (DLS) and Scanning Electron Microscopy (SEM). Fb fragment-X and β2GpI nicked at Domain-V were both prepared by limited proteolysis with plasmin [Furlan and Beck, 1972; Ohkura et al., Blood (1998)]. Domain-I was produced by solid-phase synthesis [Pozzi et al., Protein Sci (2010)].
Results: Fluorescence and SPR analyses indicate that β2GpI interacts with Fb at two sites, with Kd1 = 15±4nM and Kd2 > 1µM. Identical results were obtained with Domain-I and Domain-V clipped β2GpI. Binding of β2GpI to Fb fragment-X, lacking the Ca-domains, resulted in affinity drop. Turbidimetric, DLS, and SEM measurements indicate that 4mM β2GpI induces formation of thinner and shorter fibrin fibers.
Conclusions: Given the plasma concentrations of β2GpI (4mM) and Fb (7mM), and the affinity of β2GpI for Fb (Kd1 = 15nM), we conclude that under physiological conditions β2GpI circulates in the fibrinogen-bound form. β2GpI-Fb complex formation is driven by β2GpI Domain-I and Fb Ca-domains. Considered that Domain-I is also the major epitope for APS autoantibodies, our results challenge the current view on the role of β2GpI in the pathogenesis of APS.
To cite this abstract in AMA style:Acquasaliente L, Pagotto A, Peterle D, Pengo V, De Filippis V. β2-Glicoprotein I (β2GpI) Is a Fibrinogen Binding Protein [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/%ce%b22-glicoprotein-i-%ce%b22gpi-is-a-fibrinogen-binding-protein/. Accessed September 29, 2023.
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