Aggregation And Membrane Interaction Mechanisms Of Antimicrobial Peptides Uperin 3.5 And Uperin 3.6

Antimicrobial peptides(AMPs)have become a hot topic in the research field because of their wide spectra of antimicrobial activity.The net cationicity and amphipathicity of AMPs mediate the interaction between AMPs and anionic components of the membrane to cause cell death.In recent years,more and more researchers study the mechanism of interaction of antimicrobial peptides with membranes,but there is still no authoritative explanation for it.Uperin 3.5 and Uperin 3.6 are AMPs from the skin secretion of Australia little toads Toadlet Uperoleia mjobergii.They can inhibit gram-positive bacteria,gram-negative bacteria and fungi,thus have application potential in a lot of field beause of the antimicrobial activity.Uperin 3.5 and Uperin 3.6 share high identity in sequencewhile showed significant difference in aggregation tendency,and their antibacterial mechanisma are to be clarified yet.This study mainly investigated the aggregation differences and the revelant key sites of Uperin 3.5 and Uperin 3.6,in addition to their interaction with various membrane mimetics,in order to elucidate the relationship between their structure and aggregation and reveal the antibacterial mechanism.Firstly,using software to predict potential key sites of Uperin 3.5 and Uperin 3.6 for their aggregation,we designed mutants with these sites exchanged between Uperin 3.5 and Uperin 3.6.By thioflavin T fluorescence and circular dichroism etc,we measured the structure and aggregation of the mutants.The results show that the sites critical for the aggregation of two antimicrobial peptides are two C-terminal hydrophobic amino acids,with the aggregation difference determined by difference of the hydrophobicity of the residues.This result may contribute to clarifying the general aggregation mechanism of amyloid fibrils.Secondly,by means of thioflavin T fluorescence,circular dichroism,leakage fluorescence etc we examined the interactions of Uperin 3.5 and Uperin 3.6 with membranes of different components.The results showed that the interaction of Uperin 3.5 and Uperin 3.6 with the membranes depended on the form and component of the membrane,and the interaction had effects on the aggregation of the antimicrobial peptides and membrane integrity.These results may contribute to the clarification of the antimicrobial mechanisms of Uperin 3.5 and Uperin 3.6.