Total Chemical Synthesis and Evaluation of the Antimicrobial Properties of Porcine β-Defensin 5 Against Gram-Positive and Gram-Negative Bacteria.

Antimicrobial resistance poses a significant threat to global health. Host defense peptides (HDPs) are being investigated as a strategy to mitigate this threat. Porcine β-defensins (pBDs) are a class of HDPs present in the swine immune system. Our group previously identified pBD‑5 as a component of the innate immune system of pigs and found that a synthetically produced cysteine-protected analogue modulated gene expression in swine gut cells in response to Brachyspira hyodysenteriae infection but lacked direct antimicrobial activity. In this study, we synthesized pBD‑5 with three disulfide bonds using orthogonal protection strategies and evaluated its antimicrobial activity, structural stability at pH 7, and cytocompatibility with swine red blood cells. Circular dichroism spectroscopy revealed a secondary structure predominantly composed of β-sheets and random coils, a conformation reported for other β-defensins. Using a paired study design with three technical replicates per group, we assessed pBD-5 antimicrobial properties at eight concentrations (1.56-200 µM) against Escherichia coli and Staphylococcus epidermidis. Growth inhibition was measured by optical density (OD