Abstract
We describe the design of peptides with properties like thermostability, pH stability, and antibacterial activity against a few bacterial food pathogens. Insights obtained from classical structure-function analysis of natural peptides and their mutants through antimicrobial and enzymatic assays are used to rationally develop a set of peptides. pH and thermostability assays were performed to demonstrate robust antimicrobial activity post-treatment with high temperatures and at wide pH ranges. We have also investigated the mode of action of these hyperstable peptides using membrane permeability assays, electron microscopy, and molecular dynamics simulations. Notably, through mutational studies, we show that these peptides elicit their antibacterial action via both membrane destabilization and inhibition of intracellular trypsin—the two functions attributable to separate peptide segments. Finally, toxicity studies and food preservation assays demonstrate the safety and efficacy of the designed peptides for food preservation. Overall, the study provides a general ‘blueprint’ for the development of stable antimicrobial peptides (AMPs). Insights obtained from this work may also be combined with combinatorial methods in high-throughput studies for future development of antimicrobials for various applications.
Highlights
Microbial action is the most common cause of spoilage and one of most concern as it is associated with the risks of food poisoning and food-borne illnesses
A variety of chemical preservatives are extensively used in the food industry to improve the shelf life of food products, many of these have been associated with possible side-effects[2,3]
Analysis of natural peptides for the rational design of antibacterial properties We started with the analysis of two natural peptides: HVBBI-a β-hairpin Birk inhibitors (BBIs) found in skin secretions of Chinese bamboo odorous frog, Huia versabilis[36] and SFTI-a bicyclic trypsin inhibitor from sunflower seeds[37]
Summary
Microbial action is the most common cause of spoilage and one of most concern as it is associated with the risks of food poisoning and food-borne illnesses. It is imperative that we move toward natural and safer alternatives to address the specific needs of the area In this regard, peptide-based antimicrobials are promising due to their enhanced inhibitory activity and greater biocompatibility. Many AMPs are susceptible to cleavage by trypsin-like proteases, resulting in low efficiency at the site of action. They present the possibility of tailoring the properties and function with relative ease to achieve specific inhibition of a class of microbe. Several other groups are involved in the de novo design of antimicrobial peptides coupled with structure-function studies to develop safer alternatives to natural AMPs21–27. We attempt to design possible alternatives to antimicrobial peptides like Nisin. Some of these insights are broadly applicable to the design of antimicrobials for clinical and biomedical applications, in addition to food preservation
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