Blood: Hydrogen-thorium exchange mass spectrometry studies the conformational changes of FVIII-driven FIXa.

The assembly of the enzyme-activated factor IX (FIXa) and its cofactor activated VIII factor (FVIIIa) is a key event in the clotting cascade reaction.
loss or abnormal function of FIXa or FVIIIa can seriously affect hemorrhage and cause haemophilia.
FIXa is an inefficient enzyme that requires FVIIa to play a hemostatic role, but so far its mechanism is unclear. In a recent study published in the journal Blood, "Factor VIII-driven changes in activated Factor IX explored by Hydrogen-Deuterium eXchange Mass Spectrometry" study, Freato and others used hydroxon exchange mass spectrometry (HDX-MS) to study how FIXa responds to the assembly of FVIIIa in the presence of phospholipids.

the study reveals a complex pattern of changethat that partially overlaps with the changes that occur when the active site-oriented inhibitoroccupies the substrate binding site.
in the changes that are driven by cofactors and substrates, HDX-MS highlights several surface rings associated with the structural network of the associated clotting enzymes.
examination of FVIIIa-specific changes showed that the FIXa-FVIIIa assembly involved three spirals.
these are part of the basic interface, also known as Exosite II.
the functional study of these base residual mutations makes it clear that the interface is an extended FVIIIa interactive patch.
HDX-MS is also used in recombinant FIXa mutants associated with severe haemophilia B, suggesting that single amino acid substitution can also be used to silence FVIIIa-driven structural changes in extended networks.
in general, HDX-MS may visualize the functional effects of disease-related mutations on the function of the hemostatic system enzyme-cofactor complex.
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