Mg(II)-induced binding of factor IX-binding protein from the venom of Agkistrodon Halys Pallas with factor Xa

Factor IX-binding protein (AHP IX-bp), a Ca2+- and Zn2+-binding protein from the venom of Agkistrodon Halys Pallas was reported to bind specifically with factor IX in a Zn2+-dependent manner. Here we have purified AHP IX-bp by a simple two-step of chromatography procedure and found that AHP IX-bp also binds factor Xa (FXa) with high binding-affinity in a Mg2+-dependent manner. Although Mg2+ ions have a significantly low binding-affinity for apo-AHP IX-bp as determined by isothermal titration calorimetry, they can induce the binding of apo-AHP IX-bp with FXa even in the absence of Ca2+ as determined by native PAGE and surface plasmon resonance. Mg2+ ions are required to maintain in vivo function of FX Gla domain for its recognition of AHP IX-bp. Both Ca2+ and Zn2+ ions fail to induce the binding between apo-AHP IX-bp and FXa. The abundant Mg2+ ions in blood play an important role in the anticoagulation of AHP IX-bp.