P379 Antifungal activity of α defensins 5 like peptide against Cryptococcus neoformans var. grubii and in silico designing of its peptidomimetics

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PMObjectivesCombinatorial and retroviral therapies have been found effective in the treatment of cryptococcosis, but safer and more potent antifungal therapeutics are the need of the hour. In this regard, the natural antifungal peptides (AFPs), the essential components of the innate immune system have drawn the attention of researchers and drug developers. AFPs are 12-50 amino-acids-long host defense peptides of cationic nature. Although AFPs have certain limitations like low stability, susceptibility to proteolysis, loss of activity in the presence of metal ions and salts, short half-lives, and peptide aggregation in oral formulation, but these can be addressed by designing and synthesizing the peptidomimetics.MethodologyThe whole blood of the rat was collected by cardiac puncture. The blood was mixed with 0.83% aqueous ammonium chloride. The neutrophils were separated from lysed WBCs by centrifugation. The purification of the peptide was carried out by SDS-PAGE, size exclusion, cation exchange chromatography, and HRLCMS. The MSA of the characterized peptide was performed using a T-coffee program. The in vitro antifungal activity of crude was tested against C. n. grubii (ATCC 6352) using flow cytometry. The structure of peptides was altered through ChemDraw by replacing the elements with their bioisosteres to form mimetics. The peptide and its mimetics were docked upon the target of C. n. grubii, i.e., O-acetyl transferase (modeled through Phyre2 web server) using PatchDock, and FireDock web servers.ResultsThe peptide isolated from rat PMNs had a molecular weight of 3 kDa. It has lysine and arginine which are the characteristic residues of an antifungal peptide. The eluted peptide has the highest (83%) similarity with α defensins 5 (accession no NP_001013071 XP_214 386). The mild antifungal activity of peptide was evident against C. n. grubii. The structure of O-acetyl transferase was modeled with 100% confidence and 92% query coverage (Table 1). The mimetic 1 (M1) showed the best interaction with O-acetyl transferase with −54.47 Kcal/mol binding energy and 4 hydrogen bonds (Table 2).ConclusionThe designed M1 of α defensins 5 like peptide could be a promising lead in antifungal drug development. As mimetic 1 was designed using an in-silico approach, its synthesis and wet lab validation are desirable.