Expression and Characterization of Recombinant Triple Laterosporulin in Chlamydomonas reinhardtii

Antimicrobial peptides with broad-spectrum antibacterial properties have emerged as promising candidates for combating bacterial infections. Laterosporulin (LS), a class II bacteriocin initially isolated from Brevibacillus sp. strain GI-9, has demonstrated broad-spectrum antibacterial activity and notable biochemical characteristics. However, the limited expression levels of LS in Brevibacillus sp. GI-9 restrict its potential for large-scale production and therapeutic and food preservation applications. In this study, a triple peptide comprising three tandem repeats of LS (3 × LS), tagged with C-terminal hemagglutinin (HA) and 6 × His, was successfully expressed heterologously in Chlamydomonas reinhardtii. The purified 3 × LS-HA-6 × His effectively inhibited the growth of both Gram-positive and Gram-negative bacteria. Additionally, 3 × LS-HA-6 × His demonstrated stable antibacterial activity over a wide range of temperatures and pH values and showed significant resistance to multiple proteolytic enzymes. Furthermore, 3 × LS-HA-6 × His disrupted bacterial cell membrane integrity while exhibiting nonhemolytic activity toward rabbit erythrocytes and noncytotoxicity toward HEK293T cells. These findings underscore the potential of C. reinhardtii as a promising host for the production of bioactive and biosafe LS, providing valuable insights for the development of alternative antimicrobial strategies.