Abstract:Mytimacin-4 from Mytilus galloprovincialis has strong bacteriostatic activity against gram-positive and gram-negative bacteria. However, the production of mytimacin-4 has been rarely studied, which limits its subsequent product development and application. In this study, the gene sequence of mytimacin-4 was codon optimized in Escherichia coli and constructed in the expression vector pET-28a. Then, pET-28a-mytimacin-4 and each of the expression vectors containing different combinations of molecular chaperones (pG-KJE8, pKJE7, pGro7, pG-Tf2, and pTf16) were co-transformed into E. coli BL21 (DE3) for induced expression and activity identification. The results showed that the co-expression of molecular chaperones significantly improved the soluble expression of mytimacin-4 in Escherichia coli. Notably, the optimized yields of the recombinant strain BL21(DE3)/pET-28a-mn4/pG-KJE8 ranged from 200 to 400 mg/L, the highest reported. Purified mytimacin-4 was obtained by metal-ion affinity chromatography product of the expression products. The purified mytimacin-4 showed strong inhibitory activity against Staphylococcus aureus, E. coli, and Vibrio parahaemolyticus. In this study, an engineering strain with high expression of mytimacin-4 was constructed, which provided technical support for applying mytimacin-4 in veterinary medicine and aquatic feed.
