title: Fibrin(ogen)olytic and antiplatelet activities of a subtilisin-like protease from Solanum tuberosum (StSBTc-3).
abstract: Plant serine proteases have been widely used in food science and technology as well as in medicine. In this sense, several plant serine proteases have been proposed as potential anti-coagulants and anti-platelet agents. Previously, we have reported the purification and identification of a plant serine protease from Solanum tuberosum leaves. This potato enzyme, named as StSBTc-3, has a molecular weight of 72 kDa and it was characterized as a subtilisin like protease. In this work we determine and characterize the biochemical and medicinal properties of StSBTc-3. Results obtained show that, like the reported to other plant serine proteases, StSBTc-3 is able to degrade all chains of human fibrinogen and to produces fibrin clot lysis in a dose dependent manner. The enzyme efficiently hydrolyzes β subunit followed by partially hydrolyzed α and γ subunits of human fibrinogen. Assays performed to determine StSBTc-3 substrate specificity using oxidized insulin β-chain as substrate, show seven cleavage sites: Asn3-Gln4; Cys7-Gly8; Glu13-Ala14; Leu15-Tyr16; Tyr16-Leu17; Arg22-Gly23 and Phe25-Tyr26, all of them were previously reported for other serine proteases with fibrinogenolytic activity. The maximum StSBTc-3 fibrinogenolytic activity was determined at pH 8.0 and at 37 C. Additionally, we demonstrate that StSBTc-3 is able to inhibit platelet aggregation and is unable to exert cytotoxic activity on human erythrocytes in vitro at all concentrations assayed. These results suggest that StSBTc-3 could be evaluated as a new agent to be used in the treatment of thromboembolic disorders such as strokes, pulmonary embolism and deep vein thrombosis.
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina.
authors.forename: Alfonso
authors.initials: A
authors.lastname: Pepe
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina.
authors.forename: María Eugenia
authors.initials: ME
authors.lastname: Frey
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina.
authors.forename: Fernando
authors.initials: F
authors.lastname: Muñoz
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina.
authors.forename: María Belén
authors.initials: MB
authors.lastname: Fernández
authors.affiliation: Hospital Interzonal General de Agudos Dr. Oscar Alende, Juan B. Justo 6701, Mar del Plata, Argentina.
authors.forename: Anabela
authors.initials: A
authors.lastname: Pedraza
authors.affiliation: Hospital Interzonal General de Agudos Dr. Oscar Alende, Juan B. Justo 6701, Mar del Plata, Argentina.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Galbán
authors.affiliation: Clínica 25 de Mayo, 25 de Mayo 3267, Mar del Plata, Argentina.
authors.forename: Diana Noemí
authors.initials: DN
authors.lastname: García
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina.
authors.forename: Gustavo Raúl
authors.initials: GR
authors.lastname: Daleo
authors.affiliation: Instituto de Investigaciones Biológicas (IIB), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Universidad Nacional de Mar del Plata, Funes 3250, Mar del Plata, Argentina. Electronic address: gguevara@mdp.edu.ar.
authors.forename: María Gabriela
authors.initials: MG
authors.lastname: Guevara
mesh_terms: D001792:Blood Platelets / Q000378:metabolism; D004912:Erythrocytes / Q000378:metabolism; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000494:pharmacology; D006801:Humans; D010940:Plant Proteins / Q000737:chemistry / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000494:pharmacology; D011655:Pulmonary Embolism / Q000188:drug therapy; D011198:Solanum tuberosum / Q000201:enzymology*; D020521:Stroke / Q000188:drug therapy; D013381:Subtilisins / Q000737:chemistry / Q000494:pharmacology; D020246:Venous Thrombosis / Q000188:drug therapy
chemical_list: D005343:Fibrinolytic Agents; D010940:Plant Proteins; D010975:Platelet Aggregation Inhibitors; D013381:Subtilisins
keywords: Anticoagulant and antithrombotic agents; Haemostasis; Plant serine proteases; Platelets; Thrombolytic therapy