Abstract: Objective: The study aimed to realize the prokaryotic soluble expression of human sweat gland antibiotic DCD-1L, optimize its fermentation and purification conditions, and analyze its antibacterial properties. Methods: Firstly, the sequence of human DCD-1L was identified by searching NCBI, and the vector pET28a-DCD-1L was constructed. After transformed into E. coli ER2566, it was induced by IPTG and identified by SDS-PAGE. Secondly, through single factor experiment, the fermentation conditions of different temperature, rotation and feeding speed were studied and optimized to determine the optimal fermentation conditions for DCD-1L. Thirdly, according to the properties of the protein, a complete purification process was established, including ultrafiltration, Q-column elution and molecular sieve. Results: The optimal fermentation conditions of DCD-1L were obtained: 37 ℃, 400 r/min, and the peristaltic pump was 50 mL/30 min. The purity of DCD-1L was more than 96%, and the total recovery rate was 18.4%. The purified recombinant human DCD-1L had more significant antibacterial activity against Gram-positive bacteria such as Staphylococcus epidermidis than Escherichia coli. Conclusion: The recombinant human DCD-1L protein is expressed in prokaryotic cells, and the fermentation and purification processes is stable and reliable, which lays a foundation for the industrial production of recombinant human DCD-1L protein.