A Proteomics Analysis of Drought Stress-Responsive Proteins as Biomarker for Drought-Tolerant Sugarcane Cultivars

Antimicrobial resistance is a growing health concern. Antimicrobial peptides are a potential solution because they bypass conventional drug resistance mechanisms. Previously, we isolated a peptide from Crocodylus siamensis hemoglobin hydrolysate, which has antimicrobial activity and identified the main peptide from this mixture (QL17). The objective of this work was to evaluate and rationally modify QL17 in order to: (1) control its mechanism of action through bacterial membrane disruption; (2) improve its antimicrobial activity; and (3) ensure it has low cytotoxicity against normal eukaryotic cells. QL17 was rationally designed using physicochemical and template-based methods. These new peptide variants were assessed for: (1) their in vitro inhibition of microbial growth, (2) their cytotoxicity against normal cells, (3) their selectivity for microbes, and (4) the mode of action against bacteria using scanning electron microscopy (SEM), transmission electron microscopy (TEM) and confocal microscopy. The results indicate that all designed peptides have more potent antimicrobial efficacy than QL17 and IL15 peptides. However, only the most rationally modified peptides showed strong antimicrobial activity and minimal toxicity against normal cells. In particular, IL15.3 (hydrophobicity of 47% and net charge of + 6) was a potent antimicrobial agent (MIC = 4–12 μg/mL; MBC = 6–25 μg/mL) and displayed excellent selectivity for microbes (cf. human cells) via FACS assays. Microscopy confirmed that IL15.3 acts against bacteria by disrupting the cell membrane integrity and penetrating into the membrane. This causes the release of intracellular content into the outer environment leading to the death of bacteria. Moreover, IL15.3 can also interact with DNA suggesting it could have dual mode of action. Overall, a novel variant of QL17 is described that increases antimicrobial activity by over 1000-fold (~ 5 μg/mL MIC) and has minimal cytotoxicity. It may have applications in clinical use to treat and safeguard against bacteria.