Molecular mechanism of action of tyrocidine antimicrobial peptides using NMR spectroscopy and computational techniques

Abstract The need to come up with new and novel antibiotics that utilize unique mechanisms, to which bacteria cannot generate resistance, was the main motivation of this study. Tyrocidine peptides are non-selective antibiotics that have such properties. However, very limited information is available about their mechanism of action. The aim of this study was to determine the mechanism of action of tyrocidine peptides, tyrocidine A, tyrocidine B and tyrocidine C. Chapter 3 explored the structure of tyrocidine peptides using molecular dynamics and NMR spectroscopy. All tyrocidine peptides formed β -structures in membrane mimetic environments. Tyrocidine A and tyrocidine C formed β-structures in water, and tyrocidine B was unstructured. The results of the study of self-aggregation of tyrocidine C in water are shown in Chapter 4. Tyrocidine C monomers form dimers by coming together in sideways orientation. They could form higher ordered structures through the addition of monomers on either side of the dimer. At high concentrations, the peptide forms small aggregates, which eventually come together to form one large unstructured aggregate. The formation of higher ordered structures in water is expected to take much longer than the time simulated. iii There was very little disruption of both mammalian and bacterial membrane structures within the simulated timescales as shown in Chapter 5. However it was noted that tyrocidine peptides could permeate the membrane and possibly form aggregates that can disrupt the membrane structure. Aggregation of the peptides in the water section of the membrane reduced the interactions between the peptides and the membrane, thereby increasing the amount of time needed for the peptides to disrupt the membrane. Homology modeling based channels of tyrocidine peptides were very similar to that of gramicidin S (Chapter 6). The study of the channels in membrane indicated that they are relatively stable in the hydrophobic region of the membrane and unstable in the water phase of the membrane. These results suggested that tyrocidines could destroy bacterial cell structure through channel formation. The investigation of tyrocidine C and Ca2+ binding expected to avail an alternative non-lysis mechanism of action was difficult to ascertain. Experimental results showed that Ca2+ weakly binds tyrocidine C. This was contrary to quantum mechanical calculations, which gave plausible tyrocidine-Ca2+ complexes. The overall conclusion of this study is that, there could be several ways through which tyrocidines destroy the membrane. Events such as self-assembly, membrane binding and channel formation may all play major roles in the mechanism of action of tyrocidines.