Thrombin inhibitory peptides derived from Mytilus edulis proteins: identification, molecular docking and in silico prediction of toxicity

Optimization of the thrombin inhibitory activities of different enzymatic hydrolysates was conducted, and then an optimal hydrolysis condition by trypsin (5000 u/g) was determined as follows, digested at 45 °C and pH 8.5 for 2 h with a protein concentration of 25 mg/mL. Thrombin inhibitory activity was proved to be 76.92 ± 4.66% under this condition. A total of 39 peptides were identified in the hydrolysate by UPLC-Q-TOF–MS/MS, and all the peptides were predicted to be nontoxic by in silico predictive approaches. Twenty-six peptides were predicted to be anticoagulant peptides by molecular docking method, and the peptide 26 (Lys-Asn-Ala-Glu-Asn-Glu-Leu-Gly-Glu-Val-Thr-Val-Arg) was predicted to be a better anticoagulant peptide through both structure–activity relationship and affinity activity to thrombin. The interactional positions between peptide and thrombin were also involved in the interaction site on the S1 pocket of thrombin and strongly promoted its thrombin inhibitory activity. The firmly non-bonded interactions made the bound of peptide and thrombin firmly. Eventually, the chemical identification and activity verification of synthetic peptide 26 were conducted, and the thrombin inhibitory activity was 89.96 ± 5.30% at the concentration of 9 mg/mL. This study optimized an enzymatic hydrolysis and a virtual screening method for predicting and verifying the anticoagulant peptide from Mytilus edulis, respectively, which provided a good theoretical basis and application method for the research and development of the anticoagulant peptides, especially from the seafood products.