The human platelet antigen-1b (Pro) variant of αβ allosterically shifts the dynamic conformational equilibrium of this integrin toward the active state.

Integrins are heterodimeric cell-adhesion receptors comprising α and β subunits that transmit signals allosterically in both directions across the membrane by binding to intra- and extracellular components. The human platelet antigen-1 (HPA-1) polymorphism in αβ arises from a Leu → Pro exchange at residue 33 in the genu of the β subunit, resulting in Leu (HPA-1a) or Pro (HPA-1b) isoforms. Although clinical investigations have provided conflicting results, some studies have suggested that Pro platelets exhibit increased thrombogenicity. Under flow-dynamic conditions, the Pro variant displays prothrombotic properties, characterized by increased platelet adhesion, aggregate/thrombus formation, and outside-in signaling. However, the molecular events underlying this prothrombotic phenotype have remained elusive. As residue 33 is located >80 Å away from extracellular binding sites or transmembrane domains, we hypothesized that the Leu → Pro exchange allosterically shifts the dynamic conformational equilibrium of αβ toward an active state. Multiple microsecond-long, all-atom molecular dynamics simulations of the ectodomain of the Leu and Pro isoforms provided evidence that the Leu → Pro exchange weakens interdomain interactions at the genu and alters the structural dynamics of the integrin to a more unbent and splayed state. Using FRET analysis of fluorescent proteins fused with αβ in transfected HEK293 cells, we found that the Pro variant in its resting state displays a lower energy transfer than the Leu isoform. This finding indicated a larger spatial separation of the cytoplasmic tails in the Pro variant. Together, our results indicate that the Leu → Pro exchange allosterically shifts the dynamic conformational equilibrium of αβ to a structural state closer to the active one, promoting the fully active state and fostering the prothrombotic phenotype of Pro platelets.