Chlamydomonas reinhardtii-derived triple Oh-defensin inhibits the growth of bacteria by disrupting cell membrane integrity

Abstract

Oh-defensin, an antimicrobial peptide, extracted from the venom of Ornithoctonus hainana, potently inhibited fungi and Gram-negative bacteria but showed limited antibacterial potency against Gram-positive bacteria via an unknown mechanism. In this study, a recombinant peptide termed as 3×Oh-defensin-HA-6×His composed of three repeats of Oh-defensin followed by hemagglutinin (HA) and six histidine (6×His) double tags was expressed in Chlamydomonas reinhardtii. The bacteriostatic activity of 3×Oh-defensin-HA-6×His certified against both Gram-negative and Gram-positive bacteria was confirmed, with minimum inhibitory concentration (MIC) between 10 and 80 µg/mL. The minimum bactericidal concentration (MBC) was determined to range from 1× to 2×MIC. In addition, 3×Oh-defensin-HA-6×His exhibited resistance to temperature shocks, extreme pH, and proteinase digestion and it was biologically safe because it displayed low cytotoxicity and hemolysis. In addition, scanning electron microscopy (SEM) and propidium iodide (PI) staining revealed that 3×Oh-defensin-HA-6×His caused the destruction of the bacterial cell membrane, which effectively inhibited bacterial growth. In summary, our findings suggest that green algae-derived 3×Oh-defensin-HA-6×His holds significant promise as a potential alternative to conventional antibiotics in the future.