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Riboswitch Flip Kills Bacteria

Scientists discover a novel antibacterial molecule that targets a vital RNA regulatory element.

By Ruth Williams | September 30, 2015

Part of a riboswitchWIKIMEDIA, FDARDELResearchers at the pharmaceutical company Merck have identified a new bacteria-killing compound with an unusual target—an RNA regulatory structure called a riboswitch. The team used its drug, described in Nature today (September 30), to successfully reduce an experimental bacterial infection in mice, suggesting that the molecule could lead to the creation of a new antibiotic. Moreover, the results indicate that riboswitches—and other RNA elements—might be hitherto unappreciated targets for antibiotics and other drugs.

“Finding an antibiotic with a new target . . . has always been one of the holy grails of antibiotics discovery,” said RNA biochemist Thomas Hermann of the University of California, San Diego, who was not involved in the work. “It looks like that’s what the Merck group has now accomplished.”

The team’s research began with the idea of finding a compound that blocks the bacterial riboflavin synthesis pathway. Riboflavin is an essential nutrient for humans and bacteria alike, but while humans must consume it as part of their diet, bacteria can either scavenge riboflavin from the environment or, if supplies are lacking, make their own. “We targeted the riboflavin pathway because it is specific to bacteria so you have a built in safety margin,” said John Howe of the Merck research laboratories in Kenilworth, New Jersey, who led the research.

The team devised a simple but “very smart phenotypic screen,” said Hermann. The researchers tested roughly 57,000 antibacterial synthetic small molecules on cultures of E. coli bacteria looking for ones whose killing ability was neutralized by the presence of riboflavin. “If the effect of that antibacterial was suppressed by riboflavin,” said Howe, “then we had a good chance that the small molecule . . . was targeting the riboflavin pathway.”

The team found one molecule that fit the criteria and called it ribocil. To investigate the molecule’s mechanism of action, they applied it to cultures of E. coli cells until colonies emerged that were resistant to its effect. The researchers then sequenced the whole genomes of each of the resistant bacterial strains to find which genes were mutated.

The majority of drugs target proteins, explained Howe, “so we assumed that the mutations would be in one of the enzymes in the riboflavin synthesis pathway.” But as it turned out, while all of the 19 resistant strains did have mutations in a gene called RibB (which produces one of the riboflavin synthesis enzymes), the mutations did not affect the protein itself. They altered a non-coding part of the messenger RNA transcript: the riboswitch.

Riboswitches are regulatory elements at the beginning of messenger RNA transcripts. They bind molecules—normally metabolites—that typically suppress the transcript’s expression. “So instead of regulating the enzyme itself, [ribocil] is regulating the production of the enzyme,” Howe said.

Indeed, through reporter assays and crystallization experiments, the team confirmed that ribocil directly interacted with the RibB riboswitch, preventing expression of the protein.

“Ninety-nine-point-nine percent of drug targets are proteins,” said Hermann, “but they were prepared for the 0.1 percent outcome, and I think that’s what I really liked about this work.”

The team went on to tweak ribocil’s chemical structure, improving its killing efficiency and prolonging its effectiveness inside the body. The researchers then showed that this enhanced version of ribocil could effectively reduce bacterial burden in mice infected with a weakened E. coli strain; the bacteria are unable to efficiently expel drugs.

Weakened E. coli were used because wild-type E. coli are adept at ejecting ribocil and other compounds before they can take effect. Finding a way to keep ribocil in the bacteria and making other improvements will be necessary before it can be used as an actual antibiotic, explained Howe.

“I’ve [got] no idea if ribocil will end up being a drug candidate,” biochemist Gerry Wright of McMaster University in Ontario, Canada, wrote in an email to The Scientist, “but the work is a proof of principle, which is very important, and it makes us look to new areas of biology as targets for antibiotics.”

J.A. Howe et al., “Selective small-molecule inhibition of an RNA structural element,” Nature, doi: 10.1038/nature15542, 2015.

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Avatar of: dumbdumb

dumbdumb

Posts: 99

September 30, 2015

So, "E. coli are adept at ejecting ribocil and other compounds before they can take effect" and they have proven that bacteria can happily mutate to overcome it.

Am I missing something? Or they just proved that their research is pointless?

Avatar of: James V. Kohl

James V. Kohl

Posts: 518

October 1, 2015

Thanks for asking, dumbdumb.

They have shown that the underlying molecular mechanisms of ecological adaptation arise in the context of ecological variation in the supply of nutrients, which is linked from nutrient-dependent microRNAs to suppression of DNA damage caused by the accumulation of viral microRNAs.

Their refutation of neo-Darwinian theoretical nonsense may be interpreted by theorists to be research that is pointless because theorists like the idea of mutation-driven evolution.

Avatar of: Dennis Wert

Dennis Wert

Posts: 1

October 2, 2015

This article appears to just be a research report of possible new technique of killing virulent E. Coli and it has long way to go.  It may never be possible for this mechanism of mRNA interference to work in vivo because of the ability to adapt by wild type E. Coli,  anyway keep up the good work and remember new discoveries are always tedious e.g. Dr. Ehrlich and his magic bullet of Salvarsan - the first biochemically designed antibacterial  for syphilis prior to penicillin - he called 606 because it took 606 experiments to get the formulation and strength right.  Don't give up.

Avatar of: dumbdumb

dumbdumb

Posts: 99

Replied to a comment from James V. Kohl made on October 1, 2015

October 4, 2015

Wow!

That is one meaningless, pointless, nonsense comment.

Maybe, you are a much more deserving recipient of my nickname

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