Destabilase-lysozyme-2 — original recombinant thrombolytic preparation of medicinal leech inhibits horse platelets aggregation

Abstract

The purpose. Identifying the capacity of the medicinal leech novel original recombinant thrombolytic preparation Destabilase-Lysozyme-2 to inhibit the blood platelet aggregation. Methods. Gene of destabilase-lysozyme. ds2 (mlDL-Ds2 ), was cloned in E.coli cells. Recombinant protein was isolated in denaturing conditions using metal-chelate chromatography followed by denaturation of the polypeptide by rapid dilution in exact accordance with the procedure described by Kurdyumov A.S. et al. ( 2016, Russian Journal of Bioorganic Chemistry, v.42, s. 42–52). Blood was collected from the jugular vein of 18 horses. The functional status of platelets in the presence of different destabilase-lysozyme concentrations were evaluated for their aggregation in Platelet Rich Plasma ( PRP) and in Washed Platelet suspension (WP) using aggregometers Chrono-Log-700 and Сhrono-Log-560, USA560, США. As used aggregation inducers of ADP, collagen type III and human thrombin. Results. First demonstrated the ability of newly synthesized (Kurdyumov A.S. et al. 2016, Russian Journal of Bioorganic Chemistry, v42, s. 42–52) thrombolytic recombinant enzyme destabilase-lyzosyme to inhibit more than 40% of ADP-stimulated PRP aggregation and ADP- stimulated aggregation of horse blood washed platelets. Conclusion. The ability of destabilase-lyzosyme -2 to inhibit platelets aggregation extends biological properties of recombinant thrombolytic enzyme, pre-clinical trials which resulted in the end of 2015.