The PepSAVI‐MS Pipeline for Natural Product Bioactive Peptide Discovery

• DOI: 10.1096/fasebj.31.1_supplement.609.6
• Publication Date: 2023-11-26T17:18:44Z
• Authors: Christine L Kirkp...; David Pritchard; Nicole Parsley; Yufeng Liu; David W Hoskin; Lindsey N Shaw; Leslie M Hicks

The recent increase in multidrug‐resistant pathogens and associated morbidity/mortality demonstrate the immediate need for new antibiotic backbones with novel mechanisms of action. While natural products are a well‐studied source of biologically active small molecules, peptidyl factors contributing to their medicinal properties remain largely unexplored. To this end, we have developed the PepSAVI‐MS (Statistically‐guided bioactive peptides prioritized via mass spectrometry) pipeline to identify bioactive peptide targets from complex biological samples. To validate this pipeline, we have demonstrated successful detection and identification of a known antimicrobial peptide, cycloviolacin O2 (cyO2), from the botanical species Viola odorata. Additionally, we have widened the known antimicrobial spectrum for V. odorata cyclotides, including antibacterial activity of cyO2 against E. faecium and A. baumannii. We further demonstrate the broad applicability of the platform through the identification of novel anticancer activities for cycloviolacins by their cytotoxicity against ovarian, breast and prostate cancer cell lines. The developed platform is highly versatile as it is adaptable to any natural product source of peptides and can test against diverse physiological targets, including bacteria, fungi, viruses, protozoans, and cancer cells for which there is a developed bioassay. As such, we demonstrate extension of the pipeline to bacterial and fungal‐sourced AMPs through identification of the killer toxin KP4 from Ustilago maydis and the bacteriocin Bac‐21 from Enterococcus faecalis harboring pPD1. Using the validated pipeline, we begin to probe the vast array of natural product sources to prioritize highly active species for downstream analysis.