Identifying the biosynthetic origins of nybomycin, a reverse antibiotic

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The emergence of multi-drug antibiotic resistance in many disease-causing bacteria has raised the prospect of a devastating return to the pre-antibiotic era. It is imperative that new antibiotics are developed to counteract this problem. More than 50% of clinically used antibacterial agents are of natural product (NP) origins, with the majority produced by harmless soil bacteria called Streptomyces. We propose to study the Streptomyces natural product nybomycin.Nybomycin was first discovered in 1955 but does not dissolve in water and was not used for drug development. Very little further research was performed on until 2012 when, it was "rediscovered". Surprisingly, it was found to be active against a strain of Staphylococcus aureus with multiple resistances to antibiotics (MRSA) but not against an antibiotic sensitive Staphylococcus aureus (wild-type) strain. Further study showed that nybomycin only killed bacteria that were resistant to a class of antibiotics called quinolone antibiotics. Quinolones kill bacteria by preventing their DNA from unwinding and duplicating. This unwinding is controlled by special enzymes called topoisomerases (aka. gyrase) , and nybomycin was shown to be inactive against strains carrying normal topoisomerase genes but active against those with mutated genes resistant to quinolones.All bacteria will eventually develop resistance to whatever antibiotic they are exposed to. However, when strains of MRSA were grown in the presence of low levels of nybomycin, in order to select for nybomycin resistance, they became susceptible once again to quinolones by reversion of the relevant mutation. Due to this phenomenon nybomycin was termed a 'reverse antibiotic'. Reverse antibiotics offer the opportunity to stop the cycle of the accumulation of endless resistances, where strains of bacteria pick up more and more antibiotic resistances each time they are treated with a new antibiotic. In the case of nybomycin, it is a two-step process: bacteria resistant to quinolones are treated with nybomycin and any that develop a resistance to nybomycin are killed by quinolones.Since nybomycin is not soluble in water, it is not amenable, in its current form, to being used as an antibiotic to treat infections in humans. The aims of this research are:1. To identify the genes responsible for the production of nybomycin;2. To discover how nybomycin is made in the bacterial cell;3. To understand how nybomycin kills MRSA and other quinolone resistant bacteria.To accomplish these aims we will use the genome sequence of the bacteria which makes nybomycin and identify the genes responsible for making it. We will then delete these 'biosynthetic' and 'regulatory' genes from the bacterial genome one at a time. In some cases we will insert extra copies of the genes. After each experiment the production of nybomycin or new, related compounds, will be analysed to see which changes increase or decrease production of nybomycin or if any new compounds are produced. This will allow us to determine the sequence of events leading to nybomycin production and the function of the gene products (enzymes) involved in this process. Finally, we will study nybomycins mode of action by testing its ability to inhibit the action of various gyrase enzymes as well as the ability to kill a range of different bacteria.As part of the research we anticipate isolating nybomycin like molecules that may be starting points or the inspiration for developing an antibiotic active against MRSA.