Ribosome Rescue Inhibitors Kill Actively Growing and Nonreplicating Persister Mycobacterium tuberculosis Cells

Abstract

The emergence of Mycobacterium tuberculosis (MTB) strains that are resistant to most or all available antibiotics has created a severe problem for treating tuberculosis and has spurred a quest for new antibiotic targets. Here, we demonstrate that trans-translation is essential for growth of MTB and is a viable target for development of antituberculosis drugs. We also show that an inhibitor of trans-translation, KKL-35, is bactericidal against MTB under both aerobic and anoxic conditions. Biochemical experiments show that this compound targets helix 89 of the 23S rRNA. In silico molecular docking predicts a binding pocket for KKL-35 adjacent to the peptidyl-transfer center in a region not targeted by conventional antibiotics. Computational solvent mapping suggests that this pocket is a druggable hot spot for small molecule binding. Collectively, our findings reveal a new target for antituberculosis drug development and provide critical insight on the mechanism of antibacterial action for KKL-35 and related 1,3,4-oxadiazole benzamides.

A Small-Molecule Inhibitor of trans -Translation Synergistically Interacts with Cathelicidin Antimicrobial Peptides To Impair Survival of Staphylococcus aureus

“…1). These observations were in accord with those of E. coli and M. smegmatis (7), suggesting that the 23S rRNA target for these oxadiazoles is conserved in S. aureus.…”

“…1) (8,10). In vivo crosslinking experiments subsequently demonstrated that these oxadiazoles target 23S rRNA in E. coli and Mycobacterium smegmatis (7). To determine if the compounds have the same activity in S. aureus, we performed an intracellular photolabeling experiment using the cross-linkable oxadiazole derivative KKL-2098 ( Fig.…”

“…Cells were irradiated with UV light to activate the azide group of KKL-2098 and initiate cross-linking. The cells were lysed, total RNA was prepared, and the fluorescent reporter KKL-39703 was conjugated to the alkyne moiety of KKL-2098 using a click chemistry reaction (7). Agarose gel analysis showed a fluorescent band that comigrated with 23S rRNA from samples treated with KKL-2098, but this band was absent in the control samples treated with KKL-40 ( Fig.…”

“…1), was shown to have potent antibiotic activity against Bacillus anthracis, Mycobacterium tuberculosis, Francisella tularensis, Legionella pneumophila, and ⌬tolC strains of Escherichia coli (7)(8)(9)(10). Biochemical and cross-linking data indicated that KKL-35 and KKL-40 bind to a site on 23S rRNA that is not bound by other drugs, and this binding inhibits trans-translation but not translation (7,8). Here, we show that these oxadiazole compounds inhibit the growth of S. aureus and act synergistically with the human antimicrobial peptide LL-37 in vitro.…”

KKL-35 exhibits potent antibiotic activity againstLegionellaspecies independently of trans-translation inhibition

“…KKL-35, along with other oxadiazoles (KKL-10 and KKL-40), were identified in a high-throughput screen for inhibitors of trans-translation activity in vitro and were initially found to display antibiotic activity against several species. Oxadiazoles have since shown potent antibiotic activity against additional pathogens such as Francisella tularensis, Bacillus anthracis or Mycobacterium tuberculosis (34,42,43). The antibiotic activity of oxadiazoles on these pathogens was assumed to derive from inhibition of trans-translation but was not demonstrated.…”