Application of Fragment‐Based Screening to the Design of Inhibitors of Escherichia coli DsbA

Abstract

The thiol-disulfide oxidoreductase enzyme DsbA catalyzes the formation of disulfide bonds in the periplasm of Gram-negative bacteria. DsbA substrates include proteins involved in bacterial virulence. In the absence of DsbA, many of these proteins do not fold correctly, which renders the bacteria avirulent. Thus DsbA is a critical mediator of virulence and inhibitors may act as antivirulence agents. Biophysical screening has been employed to identify fragments that bind to DsbA from Escherichia coli. Elaboration of one of these fragments produced compounds that inhibit DsbA activity in vitro. In cell-based assays, the compounds inhibit bacterial motility, but have no effect on growth in liquid culture, which is consistent with selective inhibition of DsbA. Crystal structures of inhibitors bound to DsbA indicate that they bind adjacent to the active site. Together, the data suggest that DsbA may be amenable to the development of novel antibacterial compounds that act by inhibiting bacterial virulence.

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“…These results imply that phenamil and both BB2-50F and HM2-16F have different modes of action: both new compounds stopped rotation in phenamil-resistant stator mutants in E. coli, and in native stators in other strains, both Na + -and H + -powered. This property represents an improvement over other reported proton-powered motility inhibitors that are effective only in E. coli and require high concentrations (up to 600 µM) (42).…”

Benefits and Challenges of Antivirulence Antimicrobials at the Dawn of the Post-Antibiotic Era

“…However, the small molecule QseC inhibitor LED209 inhibited QseC signaling in homologues sharing less than 60% similarity and displayed antivirulence activity across EHEC, S. enterica and Francisella tularensis [64] constituting a promising broad-spectrum inhibitor lead. Other antivirulence targets under investigation, such as the virulence factor foldase DsbA [65], are even more diverse but can be found more widely across bacterial phyla offering exciting opportunities for designer-spectrum inhibitor development [26]. Interestingly, many important animal and plant pathogens have multiple copies of DsbA that are diverse and can be involved in specific virulence phenotypes, as for example S. enterica with three chromosomal and one plasmid-encoded DsbA copy [66], and this is something to be taken into consideration in future anti-DsbA drug selectivity assays.…”