A new twist on drug design: AdhE spirosomes as cross species anti-virulence targets

Antibiotics have been used for many years and are central to the treatment of many common bacterial infections. However, because antibiotics kill bacteria, their use increases the proportion of bacteria that are resistant to antibiotics. These resistant bacteria are naturally occurring mutants that are slightly different from the parental type of bacteria: they are not killed and are no longer susceptible to common antibiotics. This means that there is an urgent need for new alternative forms of treatment. Without effective antibiotics, the success of major surgery and cancer chemotherapy will become compromised. Also, for the NHS, the cost of health care for patients with resistant infections is higher than care for patients with non-resistant infections due to longer duration of illness, additional tests, the use of more expensive drugs and the increased need for hospitalisation.

Of all the bacteria that cause problems, the Gram-negative family are the hardest to treat because they rapidly develop resistance. One possible alternative treatment is to render the disease-causing bacteria less dangerous by turning off their offensive weapons. Compounds that "turn off' bacterial weapons are called "anti-virulence" compounds because they don't try to kill the bug, they just render them less dangerous. Imagine taking an army and removing all its key weapons- it's a lot less effective.

My lab has been developing and testing new anti-virulence compounds. These anti-virulence compounds work against several important Gram-negative bacteria, making them an exciting prospect but, to improve them, we have to know exactly how they work. In our previous work, we found a protein, called AdhE, that is a target of these anti-virulence compounds. If AdhE is deleted from bacteria, they are less able to cause disease. This tells us that AdhE is a good target against which to base and develop new treatments. However, without a protein structure, it is very hard to convert the lead compounds into drugs that are effective in treating real infections in humans.

A major breakthrough came last year when we worked with a group from Korea to solve the structure of AdhE. For the first time we are in the exciting position where we have a genuine target, its high resolution structure and lead compounds that we want to improve. We propose that solving the structure of AdhE with these lead compounds bound to the protein will allow the design of an entirely new family of anti-infective agents that could help to prevent or treat a wide range of Gram-negative pathogens. In this grant we will work across several aspects of biology and chemistry to move forward with AdhE. We are backed by strong collaborators around the world with expertise in related areas who want to help with our project.

Our recent work has focussed on characterising the bacterial acetaldehyde-CoA dehydrogenase/alcohol dehydrogenase, AdhE. Our interest in this protein was sparked by our study of the mechanism of action of a group of anti-virulence compounds that were known to down-regulate the type three secretion system (T3SS) of several pathogens. Bacteria with non-functional T3SSs are unable to deliver toxic effector molecules and are significantly attenuated or even avirulent in vivo. We found that AdhE was a target of these compounds, implying that their binding to AdhE affected the T3SS. This hypothesis was validated by deletion of the adhE gene in pathogenic E. coli O157:H7 (EHEC), as it caused a marked reduction in T3SS activity. The contribution of AdhE to EHEC virulence was tested in two in vivo systems: a rabbit intestinal model and a zebrafish infection model. In both systems, delta adhE EHEC was less able to cause disease. Moreover, the observation that AdhE may be important for virulence in several pathogens broadens its appeal as a drug target. Our hypothesis is that AdhE is an Achilles' heel for several pathogens and that compounds that block its activity can be used to block their virulence.

Through our active collaboration with Associate Prof Ji-Joon Song, we used cryo-EM to generate a 3.5 Å resolution structure of full-length AdhE from E. coli. The structure shows that the active sites of each functional domain reside on the outer and inner surfaces of the oligomeric spirosome formed by AdhE, topologically separating its two enzyme activities. For the first time, we are in the exciting position where we have a validated target, its high resolution structure and lead compounds that we want to improve. We propose that solving the structure of AdhE with lead compounds bound to the protein will allow the design of an entirely new family of anti-infective agents that could help to prevent or treat a wide range of Gram-negative pathogens.