A sodium channel protein Nav1.4-targeting polypeptide is disclosed, which has an amino acid sequence of SEQ ID NO:1. The polypeptide includes a first disulfide bond between Cys-3 and Cys-17, a second disulfide bond between Cys-4 and Cys-23, and a third disulfide bond between Cys-12 and Cys-24. The polypeptide has a complex coiled structure consisting of a main chain and disulfide bonds, in which three key residues LYS10, ARG18, and ARG22 could interact with Nav1.4 and be essential to block its penetrability of sodium. The polypeptide can target sodium channel protein Nav1.4 to form a stable protein complex; compared with naturally extracted drugs, the polypeptide Nav1.4 is superior in terms of yield, production, and cost saving, making it a promising Nav-targeting drug with broad application prospects.