The Kazal-type inhibitors infestins 1 and 4 differ in specificity but are similar in three-dimensional structure

Abstract

Blood coagulation is an important process in haemostasis, and disorders of blood coagulation can lead to an increased risk of haemorrhage and thrombosis. Coagulation is highly conserved in mammals and has been comprehensively studied in humans in the investigation of bleeding or thrombotic diseases. Some substances can act as inhibitors of blood coagulation and may affect one or multiple enzymes throughout the process. A specific thrombin inhibitor called infestin has been isolated from the midgut of the haematophagous insect Triatoma infestans. Infestin is a member of the nonclassical Kazal-type serine protease inhibitors and is composed of four domains, all of which have a short central α-helix and a small antiparallel β-sheet. Domains 1 and 4 of infestin (infestins 1 and 4) possess specific inhibitory activities. Infestin 1 inhibits thrombin, while infestin 4 is an inhibitor of factor XIIa, plasmin and factor Xa. Here, the structure determination and structural analysis of infestin 1 complexed with trypsin and of infestin 4 alone are reported. Through molecular modelling and docking, it is suggested that the protein-protein binding site is conserved in the infestin 1-thrombin complex compared with other Kazal-type inhibitors. Infestin 4 is able to bind factor XIIa, and the F9N and N11R mutants selected by phage display were shown to be more selective for factor XIIa in comparison to the wild type.

Assessment of the protein interaction between coagulation factor XII and corn trypsin inhibitor by molecular docking and biochemical validation

“…1B; Movie S1). The interactions of CTI residues Gly32 and Pro33 with the FXIIa S2 pocket are similar to interactions observed in other serine protease complex crystal structures with the chloromethylketone inhibitor PPACK [43] (which has the peptidomimetic sequence PheProArg) and protein inhibitors such as infestin-1 [44].…”

Recent Advances on Plasmin Inhibitors for the Treatment of Fibrinolysis‐Related Disorders

“…Infestins are composed of seven Kazal domains. Although native infestin did not show plasmin inhibition, recombinant infestin 1–4 (fourth to seventh Kazal domains), infestin 3–4, and infestin 4 inhibited plasmin potently with K i of 1.1, 0.4 and 2.1 nM, respectively . These Kazal domains also inhibited other serine proteases including thrombin, trypsin, factor XIIIa, and factor Xa, although not all were inhibited by each polypeptide.…”

“…Although native infestin did not show plasmin inhibition, recombinant infestin 1-4 (fourth to seventh Kazal domains), infestin 3-4, and infestin 4 inhibited plasmin potently with K i of 1.1, 0.4 and 2.1 nM, respectively. [210][211][212][213][214] These Kazal domains also inhibited other serine proteases including thrombin, trypsin, factor XIIIa, and factor Xa, although not all were inhibited by each polypeptide. Structural characterization of the seven Kazal domains revealed that domains second to fifth share high-sequence homology and thus, inhibitory properties.…”

Plasma contact factors as therapeutic targets

“…Infestin is a Kazal-type protease inhibitor isolated from the midgut of a hematophagous insect, the kissing bug Triatoma infestans . Infestin domains inhibit several coagulation proteases, with the fourth demonstrating activity against FXIIa, and some effect on plasmin and factor Xa [178]. A recombinant protein comprised of the fourth Kazal-type domain linked to human albumin reduces thrombus formation in animal venous and arterial thrombosis models, and inhibits surface induced thrombosis [179–183].…”

“…A recombinant protein comprised of the fourth Kazal-type domain linked to human albumin reduces thrombus formation in animal venous and arterial thrombosis models, and inhibits surface induced thrombosis [179–183]. Subsequent modifications have made it more selective for FXIIa than the native protein without loss of antithrombotic potency [178]. …”