Mode of action of violacein in pathogenic Gram negative bacteria and its development as an antibacterial agent

Participants: Per Sunnerhagen, Anne Farewell, Jonas Warringer, Morten Grøtli, Mate Erdelyi, Agnes Wold, Edward Moore

In the last three decades, no novel class of antibiotics targeting Gram negative bacteria has been developed. This is worrying, as resistance to one member of an antibiotics class in a bacterial population tends to confer rapid development of resistance also to other members of the same antibiotics class. Violacein is a natural compound produced by e.g. Chromobacterium violaceum, known to have antimicrobial activity against a range of bacterial species. Its investigation and further development as a candidate antimicrobial substance has been hampered by a lack of understanding of its mode of action. Violacein acts synergistically with many clinically used antibiotics, arguing that it works by a so far unexplored antibacterial mechanism.
We will use chemical genomics to identify the cellular target(s) of violacein, and thus its mode of action. This entails obtaining profiles of sensitivity for the compound to be tested (violacein) for genome-wide libraries of bacterial mutants lacking (or overexpressing) individual genes. The profiles are analyzed for clusters of mutants affected in similar functions, and from such analyses the mode(s) of action can be inferred. Candidate target proteins can then be evaluated with directed functional in vivo tests. These will be followed by in vitro testing of the violacein – target protein interaction, using biophysical methods and NMR. Knowing the mode of action, the cellular target, and exact mode of binding to that target, will make possible the chemical modification of violacein into derivatives with improved properties, such as increased solubility in water and bioavailability.
To overcome the problems associated with relatively low water solubility of violacein, we will also test alternative strategies for delivery, such as association with nanoparticles or coating with hydrophilic polymers. In addition, we will evaluate closely related compounds such as deoxyviolacein, with a more favorable toxicity profile.
In the project, we also explore other antibacterial natural and synthetic substances for their mode of action using the same methodologies, with a view to discover promising leads for further development as antibacterial agents.