Biomolecular in the bush - towards antibacterials without resistance

The RACI Division of Biomolecular Chemistry will hold its 2013 conference in the Blue Mountains in July. With themes of medicinal chemistry, chemical biology and drug discovery, here is a taste of what the meeting has in store.

Developing a new class of antibiotics that target bacterial virulence rather than viability is the focus of work by Professor Jennifer Martin, an ARC Australian Laureate Fellow based at the Institute for Molecular Bioscience at the University of Queensland (UQ).

A structural biologist, Martin’s research focuses on understanding the interactions, function and folding of medically relevant proteins and applying this to medical problems such as antibiotic resistance.

“Many antibiotics currently in use are derived from those developed in the 1950s and 1960s. Very few new classes of antibiotics have been approved in the past 20 years,” said Martin.

“These current antibiotics work by killing or inhibiting the growth of bacteria, and this puts a strong selection pressure on bacteria to develop resistance. Our research takes a different approach by targeting bacterial virulence rather than viability.”

Ancient tactics

Antibiotic resistance has been documented since the first antibiotics were identified and appears to have existed for millennia.

“Recent work analysing 30,000-year-old bacteria in permafrost showed that ancient bacteria also had antibiotic resistance genes,” said Martin.

“This is likely because many current antibiotics are derived from natural products, such as penicillin from fungus. Bacteria appear to have evolved ways of dealing with these natural products because they have been exposed to them for millennia in nature.”

Antibiotic resistance is currently developing at an alarming rate and the number of new antibiotics in the pharmaceutical development pipeline is slowing to a trickle.

The rapid rise in resistance to antibiotics is thought to be due to a number of factors including the use of antibiotics in agriculture, overuse and misuse in humans, and because bacteria readily share resistance genes with each other. These resistance genes may code, for example, for modified proteins that no longer bind the drugs or enzymes that degrade the drugs; thus, protecting the bacteria from the effects of the antibiotic.

Inhibiting protein assembly

Different classes of antibiotics exist based on their chemistry and mode of action. Most new antibiotics approved for use over the past 20 years are variations on these known classes, but they remain susceptible to the same resistance mechanisms. This is what makes Martin’s work so important.

“Our targets are the bacterial disulfide bond (DSB)-forming proteins,” said Martin, adding that her team of collaborators is one of only a few groups working in this area.

Some researchers focus on specific virulence factors produced by bacteria, such as secretion systems that are used to inject a host cell with toxins and poisonous enzymes, which helps the bacteria to enter tissues and cells and cause disease.

However, Martin’s work is different. Rather than targeting the virulence factors themselves, her team targets the enzymes that produce the virulence factors.

“This approach has merit because inhibiting the virulence factor assembly process blocks the activity of the entire arsenal of virulence factors produced by a bacteria, not just one specific virulence factor,” said Martin.

Most bacterial virulence factors have disulfide bonds and the DSB-forming proteins are required for these factors to fold and function correctly. Because virulence factors exist in the harsh extracellular environment or need to withstand host defence systems, they need extra stability: the strong covalent disulfide bond adds structural bracing to the protein to help keep it folded and functional, and to protect it from breakdown by host proteases.

Solving structures

Martin’s team is targeting the structures of two DSB proteins - the soluble DsbA protein and its integral membrane protein partner DsbB.

“Genetic knockouts of the DsbA and DsbB genes in pathogenic organisms has shown that without these proteins the organisms are no longer pathogenic,” said Martin. “We expect that by chemically inhibiting these proteins with drug-like molecules we will generate the same effect.”

Martin’s team is collaborating with Martin Scanlon at Monash University and Professors David Fairlie, Matt Cooper and Mark Schembri at UQ in the characterisation of the DsbA and DsbB proteins with the ultimate aim of developing inhibitors of these proteins. The work has a three-pronged approach to develop inhibitors.

First, they are screening libraries of drug-like fragments against DsbA proteins from human pathogens to identify potential candidates for development into drug leads.

Second, they are designing peptidomimetics. This work is based on the knowledge that the two proteins, DsbA and DsbB, interact with each other. By developing small peptide-like compounds that mimic the DsbA:B interaction, the researchers hope to block DSB activity and render bacteria avirulent.

And third, they are taking an in silico or virtual screening approach. This involves screening a virtual library of drug-like chemicals against the DSB protein structures using docking software to predict which chemicals will interact. This will enable potential DSB inhibitors to be identified via computer simulation. The best hits are then purchased or synthesised and assessed for activity in biochemical assays.

“Our focus is on Escherichia coli, at present,” said Martin, “but we are also solving the structures of DSB proteins from a number of other human pathogens to generate a library of DSB protein structures that will be available for us and the whole community to work with to develop new drugs.

“The DSB proteins appear to be excellent targets for the development of drugs that inhibit virulence rather than viability. Our goal is to generate a new chemical class of antibacterial that can be used either alone or in combination to treat infections caused by multidrug-resistant bacteria.”

Professor Jenny Martin is an ARC Australian Laureate Fellow at the University of Queensland. She has held NHMRC Senior, ARC Professorial and ARC Queen Elizabeth II Fellowships. She trained as a pharmacist in Victoria, was awarded a DPhil from Oxford University and spent her postdoctoral years at Rockefeller University. Photo: Professor Jennifer Martin.

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The Royal Australian Chemistry Institute Division of Biomolecular Chemistry will hold its 2013 conference - Biomolecular in the Bush - at the Fairmont Resort in the Blue Mountains on 14-17 July.

For more information, go to the conference website at www.raci-bio-conf.org.

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