Purification and Characterization of Trochus radiatus Derived Low Molecular Weight Bactericidal Polypeptide Active Against ESKAPE Pathogens

Antimicrobial peptides comprise core components of innate defense and act as first-line defense molecules in most marine mollusks. Marine natural products have served as templates for the design and development of novel therapeutic molecules against infectious pathogens in the past decades. In this study, we report the isolation, purification and characterization of a bactericidal peptide obtained from a marine top-snail Trochus radiatus . The crude extract of T. radiatus was assessed and found to exert potent activity against the tested fourteen gram-positive and gram-negative clinical pathogens. Three extraction buffers were used and their total protein extraction capacity from the tissue was evaluated. We then analyzed and estimated the total protein concentration and antibacterial capacity of obtained extracts. Aiming to isolate the bioactive peptide that confers antimicrobial activity, the crude extract was subjected to purification by fractional ammonium sulphate precipitation followed by chromatographic purification. The RP-HPLC and Tricine SDS-PAGE gel (15%) revealed the homogeneity of the purified antimicrobial peptide with a relative molecular mass of 7 kDa. This purified antibacterial peptide displayed potent antibacterial activity against both Gram-positive and Gram-negative bacteria’s i.e., Enterococcus faecium , Klebsiella pneumoniae and Shigella dysenteriae . The morphological changes occurred upon treatment was confirmed through electron microscopic imaging. The physicochemical characterization studies confirmed the stability of isolated peptide over variations in pH from 3 to 10, temperature from – 20 to 100 °C and in the presence of human serum. Further, the purified bioactive peptide was found to be less toxic and possess free radical scavenging activity. This is the first report of an antibacterial peptide isolated from the mollusk T. radiatus and this peptide could be developed into a peptide antibiotic alternative to the currently available synthetic or semisynthetic antibiotics.