(II) Structure and mechanistic studies of bacteriocins to further develop their potential to combat pathogenic bacteria

Bacteria have developed a number of smart tricks to combat related bacteria of the same niche by the secretion of protein antibiotics. Particularly interesting for us are bacteriocins secreted by human and plant pathogens such as Yersinia, Salmonella, Pseudomonas, Pectobacterium and E. coli. This group of bacterial antibiotics (or ‘toxins’) is highly specific in their recognition of outer membrane receptors exposed on the cell wall thereby targeting and killing closely related bacteria (for the mechanism of action see figure 1B). These natural antibiotic proteins form an important toolbox of molecules with unusual antibiotic properties including a high target (or bacterial strain) selectivity at very low minimal inhibitory concentrations (MIC in the low nM range). The high bacterial specificity of bacteriocins render this class of molecules superior to most if not all small molecule antibiotics, which typically target a broad range of bacterial species including beneficial commensal (gut) bacteria. Moreover, many of the bacteriocins target the bacterial inner membrane through pore formation and thereby undermine the rapid bacterial adaption through the formation of resistance mechanisms and hence successfully evade immunity mechanisms of bacteria.

We are primarily interested to understand the targeting, uptake and killing mechanisms of bacteriocins as the basis for future protein engineering approaches. Based on our previous studies we can e.g. engineer chimera targeting and killing Yersinia-related strains (including Y. pestis, Y. pseudotuberculosis, Y. enterocolitica and some pathogenic E. coli EHEC strains) which are also active in the presence of bacterial immunity factors. These engineered proteins could e.g. be produced by and secreted from commensal bacteria in the gut which could be given as food additives to become active species against certain infections.
In a following step we aim to introduce additional bacteriocins as highly specific antibiotics into first stages of animal and clinical research trials as a complementary way to traditional small molecule antibiotics. Initially we aim to target pathogenic Gram-negatives on the human skin such as P. aeruginosa or A. johnsonii since the application of protein-based antibiotics on the epithel has a reduced complexity and requirements e.g. regarding their stability. Because the success of this approach is primarily limited to bacteriocin producing pathogens we aim to overcome this limitation of targeting using chimera proteins engineered so that cell surface receptors of any bacterial cells not producing endogenous bacteriocins can be targeted. To maintain cellular specificity of the proteins fusions with antibody fragments, DARPins, affibodies or nanobodies raised e.g. from the Vibrio class (which do not produce bacteriocins) to be able to target a broader spectrum of pathogens.