STRUCTURE OF THE THROMBIN COMPLEX WITH TRIABIN, A LIPOCALIN-LIKE EXOSITE-BINDING INHIBITOR DERIVED FROM A TRIATOMINE BUG

Triabin, a 142-residue protein from the saliva of the blood-sucking tr iatomine bug Triatoma pallidipennis, is a potent and selective thrombi n inhibitor, Its stoichiometric complex with bovine alpha-thrombin was crystallized, and its crystal structure was solved by Patterson searc h methods and refined at 2.6-Angstrom resolution to an R value of 0.18 4, The analysis revealed that triabin is a compact one-domain molecule essentially consisting of an eight-stranded beta-barrel, The eight st rands A to H are arranged in the order A-C-B-D-E-F-G-H, with the first four strands exhibiting a hitherto unobserved up-up-down-down topolog y, Except for the B-C inversion, the triabin fold exhibits the regular up-and-down topology of lipocalins, In contrast to the typical ligand -binding lipocalins, however, the triabin barrel encloses a hydrophobi c core intersected by a unique salt-bridge cluster, Triabin interacts with thrombin exclusively via its fibrinogen-recognition exosite, Surp risingly, most of the interface interactions are hydrophobic. A promin ent exception represents thrombin's Arg-77A side chain, which extends into a hydrophobic triabin pocket forming partially buried salt bridge s with Glu-128 and Asp-135 of the inhibitor, The fully accessible acti ve site of thrombin in this complex is in agreement with its retained hydrolytic activity toward small chromogenic substrates. Impairment of thrombin's fibrinogen converting activity or of its thrombomodulin-me diated protein C activation capacity upon triabin binding is explained by usage of overlapping interaction sites of fibrinogen, thrombomodul in, and triabin on thrombin, These data demonstrate that triabin inhib its thrombin via a novel and unique mechanism that might be of interes t in the context of potential therapeutic applications.