Inhibition of Protein Secretion in Escherichia coli and Sub-MIC Effects of Arylomycin Antibiotics

Abstract

At sufficient concentrations, antibiotics effectively eradicate many bacterial infections. However, during therapy, bacteria are unavoidably exposed to lower antibiotic concentrations, and sub-MIC exposure can result in a wide variety of other effects, including the induction of virulence, which can complicate therapy, or horizontal gene transfer (HGT), which can accelerate the spread of resistance genes. Bacterial type I signal peptidase (SPase) is an essential protein that acts at the final step of the general secretory pathway. This pathway is required for the secretion of many proteins, including many required for virulence, and the arylomycins are a class of natural product antibiotics that target SPase. Here, we investigated the consequences of exposing Escherichia coli cultures to sub-MIC levels of an arylomycin. Using multidimensional protein identification technology mass spectrometry, we found that arylomycin treatment inhibits the proper extracytoplasmic localization of many proteins, both those that appear to be SPase substrates and several that do not. The identified proteins are involved in a broad range of extracytoplasmic processes and include a number of virulence factors. The effects of arylomycin on several processes required for virulence were then individually examined, and we found that, at even sub-MIC levels, the arylomycins potently inhibit flagellation, motility, biofilm formation, and the dissemination of antibiotic resistance via HGT. Thus, we conclude that the arylomycins represent promising novel therapeutics with the potential to eradicate infections while simultaneously reducing virulence and the dissemination of resistance.

Small-molecule antibiotic inhibitors of post-translational protein secretion

“…(Huang et al, 2012b). Arylomycin, with a concentration < 4 µg/mL, inhibits the extracytoplasmic localisation of 98 proteins in E. coli (Walsh et al, 2019).…”

“…(Huang et al, 2012b). Arylomycin, with a concentration < 4 μg/mL, inhibits the extracytoplasmic localisation of 98 proteins in E. coli (Walsh et al, 2019). (Pannomycin, identified in a differential sensitivity antisense assay (Parish et al, 2009), displays a non-validated inhibitory activity with an unclear mode of action.…”

In vitroeffects of sub‐inhibitory concentrations of amoxicillin on physiological responses and virulence determinants in a commensal strain ofEscherichia coli

“…However, the surviving commensals experience sub-MICs due to antibiotic clearance from the body that can exert unpredictable outcomes with time, including stimulation of horizontal gene transfer, induction of biofilm formation and spontaneous mutations, etc. incurring the risk of developing resistance (Blazquez et al 2012;Mlynek et al 2016;Gumus et al 2018;Walsh et al 2019). This increases the gravity of the situation since resistance from commensals may spread horizontally to other microbial populations in the gut.…”

Effective Small Molecule Antibacterials from a Novel Anti-Protein Secretion Screen

“…Similarly, using other assays, different screens were developed such as structure-based virtual ligand; several inhibitors for protein secretion particularly that target SecA have been discovered [ 7 , 20 , 21 , 22 , 23 , 24 , 25 ]. Sodium azide, the first known SecA inhibitor [ 26 ] is not a usable antibacterial because it is non-selective and inhibits eukaryotic enzymes such as other ATPases [ 27 ].…”