Antimicrobial Chelators and their Mechanism of Action

Abstract

Limiting the availability of metals in an environment is known to restrict bacterial growth and proliferation. For example, humans sequester metals to help prevent infection by pathogens, a system termed nutritional immunity. Chelators are small molecules that bind tightly to metals and thus have antibacterial properties that mimic these innate immune processes. This activity of chelators has not been studied extensively, although experiments with EDTA suggest that it disrupts bacterial membrane permeability by stripping lipopolysaccharide from the bacterial outer surface, possibly due to the stabilising Mg2+ and Ca2+. The work described here examines in detail the antibacterial effect of 11 chelators on Escherichia coli and how this relates to cellular starvation. Four distinct effects on cellular metal content were found with these chelators i) no change, ii) reduction to manganese, iii) reduction in zinc, and iv) reduction in iron combined with an increase in manganese. There was limited correlation between chelant metal affinities in solution with effects seen in cells. The chelants also exhibited variation in antibacterial efficacy, which was enhanced when used in combination, most yielding synergistic or additive effects. These chelants therefore offer significant potential as tools to probe metal homeostasis systems and as antibacterials.

EDTA, DTPMP and Octopirox were studied further by screening their effects on growth using a selection of E. coli mutants. DTPMP and Octopirox have similar effects on cellular metal content, depriving cells of iron and inducing uptake of manganese; mutant data suggests that DTPMP primarily affects Fe3+ uptake, while Octopirox Fe2+. All three chelators also seem t have effects on oxidative damage and tolerance, especially EDTA which deprives cells of manganese. The E. coli transcriptional response to EDTA was also investigated by RNA-SEQ. EDTA has wide-ranging effects on cellular metabolism, upregulating genes involved in carbon utilisation, energy production, translation and transcriptional regulators, including some iron-sulphur cluster proteins. Overall, the results offer the first detailed insight into the antibacterial effect of a structurally diverse group of chelants and the first step in understanding the relationship between metal affinity and their antibacterial mechanisms of action.