Characterization of thrombin inhibitory mechanism of rAaTI, a Kazal-type inhibitor from Aedes aegypti with anticoagulant activity
Autor(es): Watanabe, Renata M. O.; Tanaka-Azevedo, Anita M.; Araujo, Mariana S.; Juliano, Maria A.; Tanaka, Aparecida S.
Resumo: Saliva of blood-sucking arthropods contains a complex mixture of anti-haemostatic, anti-inflammatory and immune-modulator compounds. Among anti-haemostatic factors, there are anticoagulants, vasodilators and platelet aggregation inhibitors. Previous analyses of the sialotranscriptome of Aedes aegypti showed the potential presence of a Kazal-type serine protease inhibitor in the female salivary glands, carcass and also in the whole male, which inhibitor we named AaTI (A. aegypti thrombin inhibitor). Recently, we expressed and characterized rAaTI as a trypsin inhibitor, and its anticoagulant activity [1]. In this work we characterized the thrombin inhibition mechanism of rAaTI. Recombinant AaTI was able to prolong prothrombin time, activated partial thromboplastin time and thrombin time. In contrast, AaTI Delta (rAaTI truncated form) and C-terminal AaTI acidic tail prolong only thrombin time. In the competition assay, rAaTI, AaTI Delta or C-terminal AaTI acidic tail-thrombin interactions seem to be affected by heparin but not by hirudin, suggesting that rAaTI binds to thrombin exosite 2. Finally, the thrombin inhibition assay of rAaTI showed an uncompetitive inhibition mechanism. In conclusion, rAaTI can probably inhibit thrombin by interacting with thrombin exosite 2, and the interaction is not mediated by the AaTI C-terminal region, since the truncated AaTI Delta form also prolongs thrombin time. Research highlights: Recently, we published a paper containing original data about AaTI accepted in Biochimie. AaTI is a thrombin inhibitor expresses in salivary gland and midgut of Aedes aegypti. Our data strong suggests a different inhibition mechanism for thrombin by AaTI.
Palavras-Chave: Aedes aegypti; Kazal-type inhibitor; Midgut; Salivary gland; Thrombin inhibitor
Imprenta: Biochimie, v. 93, n. 3, p. 618-623, 2011.
Descritores: Aedes aegypti - Inflammation
Data de publicação: 2011
