Drug repurposing to target respiratory complexes of antibiotic-resistant bacterial pathogens

Antimicrobial resistance has become a global issue due to microorganisms no longer responding to antimicrobial medications. The evolutionary process of resistance has been accelerated by the misuse of antibiotics, resulting in ineffective antibiotics, leading to a global health crisis. Therefore, finding new ways to combat such a major issue has become very important, such as repurposing FDA-approved antimicrobials. Previous studies have shown that the terminal oxidase cytochrome bd may be a good drug target, due to its support for microorganisms against external stressors. The aim of this work was to repurpose the FDA-approved steroid drug quinestrol, to target cytochrome bd of the pathogen Enterococcus faecalis (E.faecalis), inhibiting its growth.

Another aim of this work was to clone E. faecalis cyd operon and transform it into an 'oxidase null' Enterococcus coli (E.coli) strain. Cloning was unsuccessful; thus, a previously cloned strain of S. aureus was used instead. The transformation was successful, and spectral work showed an unexpected result. The wild type does not produce any spectral peaks around the 620-650 range; indicative of a d-type haem, but the recombinant Staphylococcus aureus (S.aureus) cytochrome bd strain did unexpectedly produce a spectral feature within this range indicating a d-type haem. This result may suggest that antimicrobials devoid of a d-type haem, can be expressed into E. coli, which generates the modified cofactor. This is important for future research on antimicrobial resistance.

Due to unsuccessful cloning of the E.faecalis cyd operon, further work was carried out on the drug repurposing of quinestrol. Quinestrol not only inhibited growth of Gram-positive bacteria such as E. faecalis and S. aureus, but Gram-negative bacteria such as Klebsiella pneumoniae (K.pneumoniae), as well. Gram-negative bacteria have an outer membrane surrounding the peptidoglycan layer, whereas Gram-positive bacteria do not. This outer membrane can prevent drugs entering. The discovery of the effect of quinestrol on Gram-negative bacteria paves the way for further testing on this specific drug.