In-vitro antimicrobial activity of AF-DP protein and in-silico approach of cell membrane disruption.

Microbial resistance against common antibiotics has become one of the most serious threats to human health. The increasing statistics on this problem show the necessity of finding a way to deal with it. In recent years, antimicrobial peptides with unique properties and the capability of targeting a wide range of pathogens, have been considered as a potential for replacing common antibiotics. A small chitin-binding protein with anticandidal activity was isolated from Moringa oleifera seeds by Neto and colleagues in 2017, which very much resembled antimicrobial peptides. In this study, the antimicrobial protein 'AF-DP' was identified and characterized. AF-DP was heterologously expressed, purified, and characterized, and its 3D structure was predicted. Six molecular dynamic simulations were performed to investigate how the protein interacts with Gram-negative inner and outer, Gram-positive, fungal, cancerous, and normal mammalian membranes. Also, its antimicrobial and anticancer activity was assessed in vitro via minimum inhibition concentration (MIC) and MTT assays, respectively. This protein with 111 amino acids and a total net charge (of 10.5) has been predicted to be mainly composed of alpha helix and random coils. Its MIC affecting the growth of Escherichia coli, Staphylococcus aureus, and Candida albicans was 30 µg/ml, 100 µg/ml, and 100 µg/ml, respectively; AF-DP showed anticancer activity against MCF-7 breast cancer cell line. Scanning electron microscopic analysis confirmed the creation of pores and scratches on the surface of the bacterial membrane. The results of this research show that AF-DP can be a candidate for the production of new drugs as an AMP with antimicrobial activity.