X-RAY STRUCTURE OF ANTISTASIN AT 1.9 ANGSTROM RESOLUTION AND ITS MODELED COMPLEX WITH BLOOD-COAGULATION FACTOR XA

The three-dimensional structure of antistasin, a potent inhibitor of b lood coagulation factor Xa, from the Mexican leech Haementeria officin alis was determined at 1.9 Angstrom resolution by X-ray crystallograph y, The structure reveals a novel protein fold composed of two homologo us domains, each resembling the structure of hirustasin, a related 55- residue protease inhibitor, However, hirustasin has a different overal l shape than the individual antistasin domains, it contains four rathe r than two beta-strands, and does not inhibit factor Xa, The two antis tasin domains can be subdivided into two similarly sized subdomains wi th different relative orientations, Consequently, the domain shapes ar e different, the N-terminal domain being wedge-shaped and the C-termin al domain flat, Docking studies suggest that differences in domain sha pe enable the N-terminal, but not C-terminal, domain of antistasin to bind and inhibit factor Xa, even though both have a very similar react ive site, Furthermore, a putative exosite binding region could be defi ned in the N-terminal domain of antistasin, comprising residues 15-17, which is likely to interact with a cluster of positively charged resi dues on the factor Xa surface (Arg222/Lys223/Lys224). This exosite bin ding region explains the specificity and inhibitory potency of antista sin towards factor Xa, In the C-terminal domain of antistasin, these e xosite interactions are prevented due to the different overall shape o f this domain.