Structural Characteristics, Prokaryotic Expression and Activity Analysis of Antimicrobial Peptide ALFPm10 from Penaeus monodon

Antimicrobial peptides are crucial component of the invertebrates' innate immune system. Anti-lipopolysaccharide factors is a major type of AMPs in crustaceans, which can generally participate in the prevention of diseases. In this study, a new ALF named ALFPm10 was obtained by analyzing the transcriptome of hepatopancreas of Penaeus monodon. It consists of a signal peptide of 25 amino acids and a mature peptide of 98 amino acids. It contains an LBD domain and share 89% homology with ALF1 from Penaeus chinensis. The expression sequence of ALFPm10 was designed according to the amino acid sequence of its mature peptide and the preference of E. coli codon. Firstly, SUMO fusion tag was combined with ALFPm10 and connected to prokaryotic expression vector pColdIV. Then the recombinant vector pColdIV-SUMO-ALFPm10 was constructed and transformed into prokaryotic expression host strain E.coli BL21 (DE3). Induced by 0.4 mM IPTG for 9 h, the recombinant protein of SUMO-ALFPm10 was expressed in highest level and with high solubility. Moreover, ALFPm10 could be purified by affinity chromatography with the concentration of 335 mu g/mL. The sequence of ALFPm10 was conformed by LC-MS after removing SUMO tag by SUMO enzyme. Moreover, ALFPm10 at 24 mu M showed significantly antibacterial activity against six testing bacteria within 12 h. In a word, the above research would provide a great foundation to further study the functions of ALFs.