Tag Archives: Antibiotics

A ‘tail’ to tell – Lakey lab discovery could lead to a new class of antibiotics

Posted on by
Reply

 

Dr Chris Johnson

“Most of us know that we should wash our hands after being around animals but do most of us know the reasons why? As a researcher who spends most of his time in the lab killing E. coli, using E. coli specific antibiotics I should be well aware of the dangers of this often underestimated Gram-negative bacteria. However when my youngest daughter contracted E. coli O157 after visiting an agricultural show in Scotland in 2011 (even though we had followed the hand washing procedure), I realised that I did not appreciate how nasty this bacteria can be. Thankfully after 6 weeks in hospital including a lengthy stretch on dialysis my daughter made a full recovery but not everyone is so lucky.”

Professor Jeremy Lakey

The urgent need to develop new drugs to target pathogenic bacteria has been a theme of the ICaMB blog since its inception in early 2013.  However, these words from Dr Chris Johnson, a postdoc in Jeremy Lakey’s laboratory, bring home the seriousness of the problem.  Fortunately, Chris is in a position to do something about this.

Many of you will have realised that last week saw another potential breakthrough in ICaMB’s ongoing ‘War on bacteria’ (and here, here and here).   This time the PI making the news, with data demonstrating the possibility of a whole new class of antibiotics, was Professor Jeremy Lakey.  And when we say making the news, we mean that literally. The ITV interview of Jeremy and his team can be viewed here

This story has made news around the world and has been featured in newspapers in Australia and India (and here), as well as closer to home.  Here is the official university press release.

So what’s going on?  If you want to read the primary paper it is here

The unstructured domain of colicin N kills Escherichia coli. Mol Micro 89:84-95

Chris Johnson, who was the lead author on this manuscript explains:

“E. coli produces protein antibiotics called colicins which are used to kill E. coli and closely  related bacteria in the eternal bacterial arms race. In order to further understand how one of these, colicin N (ColN), works, we dissected the protein into its individual domains to see how each part behaved in isolation. Quite by chance we found that part of the protein, THE TAIL, was actually toxic to them. Although far less efficient than the entire ColN molecule, it remains specific for E. coli and furthermore, the specificity is housed within an intrinsically unfolded domain (a domain which has no defined 3D structure). Although this is a very basic discovery in its early stages, it allows us to appreciate novel mechanisms to kill E. coli.” (see the full version of this at the bottom of the page)

Some more about Jeremy

Any of you who were bioscience undergraduates in Newcastle will know Jeremy Lakey from his famous recreations of protein structure using party balloons.  Others may know him for co-founding Orla Protein technologies.  Some of us know him as a man who will always buy his round in the pub.  All of us know him as a great scientist and colleague.  Furthermore, Jeremy is a leading supporter of Leading Edge and recently a group of 6 Year 9 school pupils from St Cuthberts RC School looked at how ColN acts against E.coli when you start changing the amount of salt they are grown in. As many ICaMB scientists may know Jeremy also runs a workshop with Ponteland Community High School to explore bacterial shape and their surfaces using LEGO.

But did any of us think that one day Jeremy Lakey may SAVE THE WORLD from antibiotic resistant bacteria? Possibly.

The detailed science

Most Gram- negative bacteria produce protein antibiotics which are used as weapons in the battle between competing populations of bacteria.  E. coli produces protein antibiotics called colicins which are use to kill E. coli and closely related bacteria.  Once colicins are released into the extracellular milleu they dock onto their targets via specific outer-membrane receptors and then seek out an internal, periplasmic, binding partner (the Tol or Ton proteins) which helps them translocate into the cell.  We study colicin N (ColN) which comprises of an intrinsically unfolded N-terminal translocation (T) domain, involved in TolA and OmpF binding.  Its central receptor binding (R) domain binds lipopolysaccharide whilst its C-terminal 200 amino acids define the cytotoxic pore-forming (P) domain. This latter feature is common to all pore-forming colicins and forms a channel in the inner-membrane causing K+ release and cell death.  Other colicins have C-terminal domains which display cytotoxic activities that include DNAase or RNase activity.  Irrespective of the particular cytotoxic activity, all colicins are comprised of three domains (T-R-P) and it was assumed that the sole role of the T and R domains was to deliver the cytotoxic C-terminal domain across the outer-membrane.

In order to investigate the mechanism of ColN activity we dissected the protein into its individual domains.  We were attempting to block the toxic activity of full length ColN by pre- incubating E. coli cells with the intrinsically unfolded T-domain.  The rationale behind the experiment was that we could block all the available receptor sites on the E. coli target cells by saturating with T-domain, such that when the full length ColN was added to cells it would be non-toxic, as all the essential receptor binding sites would be already sequestered.  However rather than protecting the cells, T-domain was found to be toxic and like full length ColN provoked K+ efflux.   Although less efficient than full length ColN, T-domain is strictly dependent upon the same receptor proteins, OmpF and TolA for killing.  Since these receptors are only found in E. coli-like bacteria, T-domain displays the unusual combination of a generic killing mechanism coupled with extreme specificity housed within an intrinsically unfolded domain.

Links

Jeremy Lakey’s University home page: http://www.ncl.ac.uk/camb/staff/profile/jeremy.lakey

Follow Jeremy Lakey on Twitter: https://twitter.com/JeremyLakey

ICaMB: http://www.ncl.ac.uk/camb/

The Centre for Bacterial cell Biology: http://www.ncl.ac.uk/cbcb/

The official Newcastle University press release: http://www.ncl.ac.uk/press.office/press.release/item/chance-finding-could-lead-to-new-antibiotics

Link to ITV story:

http://www.itv.com/news/tyne-tees/story/2013-07-05/breakthrough-in-combatting-bacterial-infection/

The ‘Australian’ story: http://www.theaustralian.com.au/news/breaking-news/tail-could-be-used-for-new-drugs/story-fn3dxix6-1226674131684

The ‘Times of India’ Story: http://articles.timesofindia.indiatimes.com/2013-07-06/science/40406695_1_escherichia-coli-e-coli-protein

The Northern Echo story: http://www.thenorthernecho.co.uk/news/10524003.Scientists__chance_find_may_develop_new_generation_of_antibiotics/

Jeremy’s company, Orla Proteins: http://www.orlaproteins.com/about-orla/the-board.aspx

 

Science Minister visits Centre for Bacterial Cell Biology

Posted on by
Reply

 

by Dr Heath Murray 

On June 27 the RH David Willetts MP, Minister for Universities and Science, visited the Centre for Bacterial Cell Biology (CBCB) to hear about how research on bacteria can lead to: development of novel antibiotics, design of synthetic biological systems, and even understanding the origins of life on earth. Dr Heath Murray (CBCB & ICaMB) tells us more about this visit.

Mr. Willetts was given a guided tour of the new Baddiley-Clark building by the director of the CBCB, Prof Jeff Errington.

Jeff (left) outlines CBCB research to David Willetts (right), with Heath (middle back) paying close attention

Jeff discussed why he left Oxford University after 25 years to start the CBCB at Newcastle, the first Centre of its kind in the UK to provide a world-class facility in which to carry out fundamental research on bacterial cells. During the tour Jeff highlighted how the localised network of international researchers at the CBCB, working on biological problems in model bacterial organisms provides an unparalleled setting in which to exchange ideas and to benefit from related advances in microbial cell biology. While walking around Jeff noted how the open plan of the Baddiley-Clark building promoted interactions amongst the various research groups, thereby creating a uniquely stimulating environment for the scientists that work there.

This was a very fruitful visit with interesting discussions, as highlighted by Jeff: “I was impressed at how quickly the Minister picked up the key biological points we wanted to make, such as about how our work impacts on thinking about the origins of life!”

An image similar to those seen by David Willetts showing severe DNA segregation defect in a mutant Bacillus subtilis strain, observed using epifluorescent microscopy. (DNA: blue; origin of replication: green, cell membrane: red)

 

I then demonstrated the bespoke microscopes available within the CBCB to the Minister, highlighting how the small size of bacterial cells (only a few micrometers) makes microscopic analysis technically challenging and how the CBCB is utilizing state-of-the-art super-resolution microscopes to overcome this difficulty. I also explained how researchers use genetic engineering to fuse their “proteins of interest” to the Green Fluorescent Protein (GFP) from the jellyfish Aequorea victoria, thus creating tools to visualize the localization of proteins or nucleic acids within living bacterial cells using fluorescence microscopy.

 

Heath explains the potential applications of the research to the Science Minister

 

The Minister was keen to see the live demonstration of our fluorescent microscope and seemed amazed by how clearly the organization of the bacterial chromosomes was immediately apparent. He quickly appreciated that interfering with this process might have application in the development of new antibiotics.

 

 

We were all left with the clear feeling that Mr. Willetts enjoyed hearing about the science taking place within the CBCB and how this fundamental research provides insights crucial for the discovery and development of new antibiotics, as well as providing solutions to a wide range of industrial and environmental problems. “It was an interesting meeting – very reassuring to hear that the Minister is keen to make sure that Government continues to invest in Blue-Skies Research”, Jeff concluded.

 

Spills and pills: thrills for a structural biologist

Posted on by
Reply

One of the newest recruits to ICaMB is Professor Bert van den Berg, who arrived here in December 2012.  Bert is already off to a great start having been awarded a Royal Society Wolfson Research Merit Award in April.  Here we have asked him to tell us why he decided to join ICaMB and the research that lead up to this prestigious award.

By Bert van den Berg


Bert, looking thrilled

I joined ICaMB in January, coming from the University of Massachusetts Medical School in Worcester, where I was a tenured faculty member in the Program in Molecular Medicine. While I had a great and productive time in this department, after eight years I felt increasingly isolated academically and started to look for another position. ICaMB seemed a great fit for my research interests, with a large number of scientists interested in bacterial biochemistry and cell biology. Since ICaMB was also looking to strengthen its efforts in structural biology, the decision to cross the pond and join ICaMB wasn’t a very hard one. I am happy to be here, and I hope and expect that my expertise in membrane protein structural biology will also be a benefit for the faculty within ICaMB and will lead to successful collaborations.

My lab has been studying protein channels (see below) for about nine years. Determining structures is really the only way to obtain deep insights into protein function. In addition, seeing a new protein structure for the first time is often an “aha!” moment and, at least for me, the closest thing to a true discovery in modern science. In any case, the importance of structural biology for science is clear from the large number of Nobel prizes awarded to the field over the years.

What do the cleanup of oil spills and the treatment of many bacterial infections have in common? The answer is that both processes depend on the efficient passage of bacterial membranes by small molecules.

Oil spills and antibiotics have more in common than you may realise

Gram-negative bacteria are surrounded by two lipid membranes, which are termed plasma membrane and outer membrane. The outer membrane borders the cell and is a very efficient and sturdy barrier that protects the cell from noxious substances in the external environment, such as bile acids in the case of E. coli bacteria living in the gut. However, since bacteria also require nutrients for growth and function, protein channels are present in the outer membrane to allow the uptake of such small molecules. In our work we use X-ray crystallography to determine the atomic 3D structures of the channels, most of which are shaped like hollow barrels. Based on the structures we propose transport models, which we then test by characterisation of mutant proteins.

Many Gram-negative bacteria are able to use industrial pollutants such as oil as food sources, a process called biodegradation. The enzymes that catalyse these remarkable processes are located inside the cell but not much is known about how the pollutants enter the cell in the first place, something that is clearly required before they can be degraded. We study the highly specialised channels that mediate the uptake of these water-insoluble (“hydrophobic”) molecules. In addition, we are interested in discovering cellular adaptations that allow biodegrading bacteria to grow on these toxic compounds. We think that this research may lead to insights that will aid the design of bacterial strains that are optimised not only for bioremediation but also for important other processes such as production of biofuels.

The other main focus of research in my lab is to understand how antibiotics “hijack” outer membrane channels to enter bacteria. Being water-soluble, antibiotics are dependent on protein channels for membrane passage. Bacteria that are under antibiotic pressure will often change or remove the channels through which antibiotics pass, resulting in resistance.

Movie showing ampicillin movement through E coli OmpF protein channel. The view is from the outside of the cell. Movie made by Matteo Ceccarelli (University of Cagliari).

In concert with other mechanisms such as enzymatic degradation and increased efflux by pumps, this acquired antibiotic resistance has the potential to become a huge and global problem in public health. New drugs are therefore urgently needed. The problem is that not nearly enough new drugs are currently in pharmaceutical pipelines, due to the costly and risky nature of antibiotic development. However, pharmaceutical companies are starting to realise that the fundamentals of drug design need to change, and that they have to collaborate with academic labs that are studying the basic biology of small molecule membrane transport.

My lab is participating in an exciting, EU-funded joint venture between big pharma, small biotech firms and academic labs aiming to understand the influx/efflux of drugs in a number of pathogenic Gram-negative bacteria. Beyond the potential benefits for drug design, it is hoped that this project will change the way in which industry and academia work together to benefit public health.

 

Links

Royal Society Wolfson Merit Awards: http://royalsociety.org/news/2013/new-wolfson-research-merit-awards/

Bert’s ICaMB homepage: http://www.ncl.ac.uk/camb/staff/profile/bert.van-den-berg

Newcastle Structural Biology website: http://sbl.ncl.ac.uk/people/bert_research.shtml

Structural Biologist Nobel Prize Winners: http://www.ebi.ac.uk/pdbe/docs/nobel/nobels.html

IMI TRANSLOCATION project: http://www.imi.europa.eu/content/translocation