Separation and Identification of Novel Antithrombotic Protein and Peptides from Shrimp Paste Hydrolysates

Antithrombotic activity is primarily manifested in two aspects: anticoagulation and thrombolysis. It has been observed that shrimp paste has notable antithrombotic activity. However, research on mining active substances from shrimp paste combining anticoagulation and thrombolysis is limited. This study optimized Litopenaeus vannamei shrimp paste via enzymatic-assisted fermentation to enhance its dual antithrombotic profile and identify the responsible components. Optimal activity was achieved by using 1% chitosanase at 45 °C and pH 7.0 for 6 h, followed by fermentation at 25 °C for 15 d. A fibrinolytic fragment (approximately 11.36 kDa) from the SIT-AKD region of protein A0A423T5F8 showed activity equivalent to 40.28 μg urokinase at 21 μg. Meanwhile, peptides PSEPSKPVTCKPR and GIAEGCDYPWR exhibited 1.35-fold and 1.19-fold higher anticoagulant activity than heparin sodium at 3 mg/mL. Molecular docking indicated that hydrogen-bonding interactions with Trp60, Glu192, Gly216, and Arg221 underlie the anticoagulant mechanism. These findings support the high-value utilization of shrimp paste.