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Research article2003Peer reviewed

Heparin and calcium ions dramatically enhance antithrombin reactivity with factor IXa by generating new interaction exosites

Bedsted T, Swanson R, Chuang YJ, Bock PE, Bjork I, Olson ST

Abstract

Blood coagulation factor IXa has been presumed to be regulated by the serpin, antithrombin, and its polysaccharide activator, heparin, but it has not been clear whether factor IXa is inhibited by the serpin with a specificity comparable to that for thrombin and factor Xa or what determinants govern this specificity. Here we show that antithrombin is essentially unreactive with factor IXa in the absence of heparin (k(ass) similar to 10 M-1 s(-1)) but undergoes a remarkable similar to1 million-fold enhancement in reactivity with this proteinase to the physiologically relevant range (k(ass) similar to 10(7) M-1 s(-1)) when activated by heparin in the presence of physiologic levels of calcium. This rate enhancement is shown to derive from three sources: (i) allosteric activation of antithrombin by a sequence-specific heparin pentasaccharide (300-500-fold), (ii) allosteric activation of factor IXa by calcium ions (4-8-fold), and (iii) heparin bridging of antithrombin and factor IXa augmented by calcium ions (130-1000-fold depending on heparin chain length). Mutagenesis of P6-P3' reactive loop residues of antithrombin further reveals that the reactivity of the unactivated inhibitor is principally determined by the P1 Arg residue, whereas exosites outside the loop which are present on the activated serpin and on heparin are responsible for heparin enhancement of this reactivity. These results together with our previous findings demonstrate that exosites are responsible for the unusual specificity of antithrombin and heparin for three clotting proteases with quite distinct substrate specificities

Published in

Biochemistry
2003, Volume: 42, number: 27, pages: 8143-8152
Publisher: AMER CHEMICAL SOC

      SLU Authors

    • Björk, Ingemar

      • Department of Molecular Biosciences, Swedish University of Agricultural Sciences

    Publication identifier

    DOI: https://doi.org/10.1021/bi034363y

    Permanent link to this page (URI)

    https://res.slu.se/id/publ/1085