Abstract
ABSTRACTHuman health is threatened by bacterial infections that are increasingly resistant to multiple drugs. A recently emerged strategy consists of disarming pathogenic bacteria by targeting and blocking their virulence factors. The type VI secretion system (T6SS) is a widespread secretion nanomachine encoded and employed by pathogenic strains to establish their virulence process during host invasion. Given the conservation of T6SS in several human bacterial pathogens, the discovery of an effective broad-spectrum T6SS virulence blocker represents an attractive target for development of antivirulence therapies. Here, we identified and validated a protein-protein interaction interface, TssK-TssG, as a key factor in the assembly of the T6SS baseplate (BP) complex in the pathogen enteroaggregative Escherichia coli (EAEC). In silico and biochemical studies revealed that the determinants of the interface are broadly conserved among pathogenic species, suggesting a role for this interface as a target for T6SS inhibition. Based on the high-resolution structure of the TssKFGE wedge complex, we rationally designed a biomimetic cyclic peptide (BCP) that blocks the assembly of the EAEC BP complex and inhibits the function of T6SS in bacterial cultures. Our BCP is the first compound completely designed from prior structural knowledge with anti-T6SS activity that can be used as a model to target human pathogens.
Highlights
IMPORTANCE New therapeutic options are urgently needed to fight drug-resistant and life-threatening infections
None of them is efficient against the Gram-negative WHO priority list ESKAPE pathogens, namely, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter species, which are responsible for approximately 75% of infections and deaths by antibiotic-resistant bacteria [2, 3]
In order to diagnose at which stage the T6SS biogenesis was affected by TssG foot deletion, we biochemically analyzed the behavior of the TssKFGE unit using native gel experiments and observed that deletion of either foot 1 or foot 2 destabilizes the wedge complex (Fig. 1C and Fig. S1A and C)
Summary
IMPORTANCE New therapeutic options are urgently needed to fight drug-resistant and life-threatening infections. None of them is efficient against the Gram-negative WHO priority list ESKAPE pathogens, namely, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacter species, which are responsible for approximately 75% of infections and deaths by antibiotic-resistant bacteria [2, 3] In this postantibiotic era, new therapeutic options are required to fight against drug-resistant and life-threatening infections. The high-resolution structure of the TssKFG complex from the pathogen enteroaggregative Escherichia coli (EAEC) has been solved and provided incisive insights into the structural organization of the protein complex [50, 51] This 500-kDa complex is composed of a monomeric TssG serving as a central backbone that interacts with a TssF dimer and two TssK trimers. Targeting the T6SS Baseplate by a Biomimetic Peptide truncated version of TssG lacking foot 1 and foot 2 interferes with baseplate assembly and leads to T6SS function impairment [50]
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