Novel Peptide Motifs Containing Asp-Glu-Gly Target P2Y12 and Thromboxane A2 Receptors to Inhibit Platelet Aggregation and Thrombus Formation

Increasing evidence has shown that collagen peptides have multiple biological activities. Our previous study has separated and identified antiplatelet aggregation peptides Asp-Glu-Gly-Pro (DEGP) from Salmo salar skin. This study is to investigate the cellular target of DEGP on platelets and its underlying mechanism. DEGP inhibited platelet aggregation in a dose-dependent manner induced by 2MeS-ADP and U46619 and significantly attenuated tail thrombosis formation by 30% in mice at the dose of 50 mg/kg body weight. Mechanically, DEGP displayed apparent antagonism effects on TP and P2Y12 receptors by the drug affinity responsive target stability (DARTS) technique to regulate the phosphorylation of RhoAS188, PLCβ3S537, as well as VASPS157. The molecular docking results revealed a stronger binding energy with the target protein of modified peptides DEGI and DDEGL. Practically, DEGI exhibited the highest inhibition activity against 2MeS-ADP- and U46619-induced platelet aggregation in vitro with IC50 values of 0.88 ± 0.10 and 0.85 ± 0.10 mM, respectively, and comparable antithrombosis activity with aspirin at the dose of 25 mg/kg body weight in vivo. These results indicated the possibility that the peptide motifs containing Asp-Glu-Gly could potentially be developed as a novel therapeutic agent in the prevention and treatment of thrombotic diseases.