Paper of the month
We have selected one scientific publication per month, to provide a flavour of the variety of research going on in the Department of Molecular Biology and Biotechnology. Further information can be found in the links.
July 2017 - A gene that links motor neurone disease and dementia
Research published by Prof Sherif El-Khamisy's group and researchers in Sheffield's Medical School in the journal Nature Neuroscience (July 17) has identified a novel mechanism of disease, and a way of stopping it. They studied a gene called C9orf72, which is known to be linked to motor neurone disease (MND) and frontotemporal dementia, and showed that there is a 6-base sequence in its DNA that can be duplicated multiple times. When this happens, the DNA makes structures called R-loops which are easily damaged. Cells have a mechanism for getting rid of damaged DNA, but in this case this mechanism over-activates the cell's 'garbage bin' system, called autophagy (from the greek words for 'eating yourself'), which means that the cells get killed off, leading to neurological damage. The researchers showed that if they turned up the cells' natural DNA repair systems, then the cells no longer needed to over-activate autophagy, and cells were able to cope. This is a potential treatment for devastating diseases such as MND, which is one of the most common neurodegenerative disorders affecting younger people in the middle of their active life.
June 2017 - Why stomata in plant leaves are so important
Stomata are the holes in plant leaves that let water out (and hence support transpiration, which sucks water up from the roots) and also let carbon dioxide in (to provide carbon for photosynthesis). The picture shows a typical example, from the model plant Arabisopsis. As global warming increases, it would seem sensible to reduce the number of stomata, because there is more carbon dioxide in the atmosphere and a greater need to conserve water. A series of three papers from Prof Julie Gray in the journal Plant Physiology has shown that this is a feasible solution. In the first paper, she shows that stomata evolved very early on in land plants and are well conserved, implying that ideas based on one plant should be transferable to others. In the second, she describes the structure of their outer edge. And in the third, she proves the key hypothesis by showing that when the number of stomata in barley are reduced, they show greater tolerance to drought without impact on yield.
May 2017 - A view of a bacteral spore outer coat
Some bacteria have a protective coat around them, called the exosporium. This includes the genera Bacillus and Clostridium, such as Bacillus anthracis (responsible for anthrax) and Clostridium difficile, the hospital superbug. The coat protects the bacterial spores and enables them to survive for years in a dormant state; it also helps to attach them to a host. Profs Per Bullough and Anne Moir have combined forces to work out how the exosporium is made. They have used molecular biology to work out the protein components of the coat, and electron microscopy to work out where they are and how they fit together. The picture shows a folded piece of exosporium from the plant pathogen Bacillus thuringiensis at high magnification (the bar represents 100 nm). In a paper in Molecular Microbiology (Vol 104 page 539) they show how the exosporium is self-assembled into a two-dimensional crystalline array. The challenge now is to use this detailed information to find novel ways of attacking these pathogenic bacteria.
April 2017 - DNA strand repair in mitochondria
We have known for a long time that correct repair of damaged DNA is critical for cell viability. Mitochondria (the powerhouses of cells) have their own DNA, and it has always been assumed that they similarly need to repair damaged DNA - in fact even more so, because several of the chemical reactions that take place in mitochondria generate free radicals, which are very damaging to DNA. However, until now we have not known how they do it. Research published by Prof Sherif El-Khamisy has now shown the key role played by the enzyme tyrosyl DNA phosphodiesterase 1. For more on this story, visit Science Advances April 2017 vol 3 issue 4, page e1602506. There is a short video on this at that site, which you can also find on our News web page.
March 2017 - How phosphoryl transfer enzymes work
You might think that we understand how enzymes manage to accelerate reaction rates so much, and so specifically. However, this is not true - for most enzymes we only have a vague idea how they work. Enzymes that catalyse phosphoryl transfer include kinases, ATPases and phosphatases, and therefore constitute arguably the most important group of enzymes. They increase the reaction rate by up to 1020-fold - a truly remarkable rate increase. Studies by Profs Jon Waltho and Mike Blackburn, reported in Angewandte Chemie 2017 56:4110, use a combination of X-ray crystallography, NMR, and computational analysis, using metal fluorides as analogues of the phosphate group. They show how these enzymes use a combination of precise geometrical positioning, charge balance, and control of hydrogen bonds, to achieve their rate enhancements. They also show how much we still have to learn.
February 2017 - Improving light capture for photosynthesis
Photosynthetic organisms do not use all of the light energy they receive from the sun, because their pigments (such as chlorophyll) only absorb light at selected wavelengths. Prof Hunter and his team have come up with a way to capture a larger range of wavelengths, and to channel this energy into the bacterial photosynthetic reaction centre so that the energy can be used for photosynthesis. They took the gene that encodes the photosynthetic reaction centre, and fused it to the gene for yellow fluorescent protein (YFP). YFP absorbs green light (not used by the photosynthetic bacteria) and re-emits it as yellow, which the bacteria can absorb. This led to more energy capture and faster photosynthetic growth. The figure shows the structure of the photosynthetic centre they produced. The large circle is the normal photosynthetic reaction centre, and the smaller circle at the side is YFP, which is close enough to pass its energy to the reaction centre via the pigment proteins in blue. This work was published in Nature Communications 2017 8:13972.
January 2017 - Biogenesis of peroxisomes
Peroxisomes are organelles found in almost all mammalian cells. Their best understood functions are in breaking down long chain fatty acids, but they have many other functions too, and genetic diseases involving loss of peroxisome function create a range of problems, particularly in the nervous system. Dr Hettema has been studying how peroxisomes develop. In most cases, new peroxisomes bud off existing ones. But they can also appear in cells that have no peroxisomes at all - so where do they come from? In a paper published in Nature, Dr Hettema discusses a recent result showing that they can be created from membrane pinched off from other existing internal membranes - mitochondria and endoplamic reticulum (ER). This is not yet a complete answer to the question, but provides an important step along the way.
December 2016 - Exposing weaknesses in the bacterial cell wall to prevent infection
It is becoming critically urgent to find new ways of killing bacteria, to get round the problem of bacterial resistance. New lecturer Dr Fenton has just found a completely new target, which was identified in Streptococcus pneumoniae (which causes pneumonia and meningitis) but seems to be widespread in bacteria. This is a control system, which regulates when and where bacteria insert a critical component called lipid II into their cell walls. Disruption of the control system leads to lipid II being inserted all over the bacterial coat rather than in the desired places, as a result of which the bacteria burst apart and die. The aim now is to work out how to disrupt the control most effectively. The work was published in Nature Microbiology, doi:10.1038/nmicrobiol.2016.237.
November 2016 - Predicting protein solubility and stability
Many proteins are not very soluble, and many are not very stable in solution - they precipitate. This is a problem for researchers trying to study the proteins, and also a problem for pharmaceutical and life science companies trying to store and use proteins. Getting proteins to behave better in solution is a black art. About the only rule is the Hofmeister series: ions at one end of the series (like sulphate) stabilise proteins but make them less soluble, while ions at the other end (like thiocyanate) destabilise proteins but make them more soluble. There are conflicting theories to explain this effect. Work published by Prof Williamson and colleagues in the American Chemical Society open access journal Omega shows that the effect is due to how the ions interact with water. Sulphate orders water around it, and therefore makes the water molecules less free to solvate proteins. This understanding should lead to new approaches to stabilise and solubilise proteins - and indeed Prof Williamson has been awarded a research grant starting in 2017 to help him develop this.
October 2016 - How to stop cancer cells developing drug resistance
Most chemotherapy and radiotherapy kill cancer cells by causing breaks in their DNA. Unfortunately, cancer cells fight back by fixing the breaks through a specialised toolkit, causing cancer resistance. If we could find a way to hijack the cancer’s fix and repair toolkit and make it less efficient, then we could tip the scale in favour of cancer cell death instead of survival (i.e. resistance).
Research published by Prof El-Khamisy and Dr Hodgson in MBB and others in the journal Nucleic Acids Research (27 Oct 2016) showed that resistance to a common class of chemotherapy used to treat colon and breast cancer is caused by changes in the “speed" by which the tool kit components travel to, and stay at, the sites of DNA breaks. They were able to identify a change in a specific mark in the proteins that wrap the DNA – called histones – in such a way that makes repair much faster in cancer cells (i.e. make them resistant to therapy). This means that the change in cancer cells is not genetic but epigenetic. As a result of this discovery, the group worked out a way to inhibit the activity of the enzyme that puts in the histone marks, thus resetting the epigenetic change and reversing the resistance of the cancer cells. This provides a new way of stopping cancer cells developing drug resistance.
September 2016 - Design of a herbicide drug
Nearly all chiral drugs are produced as single enantiomers - just look up Thalidomide on the internet to see why. It was therefore a surprise when a Sheffield team led by Prof Rice and Dr Baker from MBB, in collaboration with the plant protection group at Syngenta, crystallised a novel inhibitor of IGPD (an enzyme involved in the biosynthesis of histidine, and a good target for a novel herbicide to replace Glyphosate when weeds eventually become resistant to it) bound to the target enzyme. They found that both enantiomers of the inhibitor bound equally tightly, with both fitting well into the active site of the enzyme though with opposite orientations (coloured central figure). It turned out that they are mimicking the Δ2-enol intermediate of the reaction, which is flat. This was a surprise because the inhibitor C348 (bottom right) was designed to mimic the diazafulvene intermediate, which is not flat. This finding provides new opportunities for improving the design of inhibitors. The work was published in the top Chemistry journal Angewandte Chemie Int Ed (Sept 26).
August 2016 - Growth and processing of algae for biofuels
A promising source of sustainable biofuels is algae, which can be selected and grown for efficient production of lipids, useful for high-grade biofuels. However, economic use of algae relies on efficient conditions for growing them, and subsequently extracting the lipids. Research from Dr Jim Gilmour in MBB, in collaboration with colleagues in Chemical and Biological Engineering, has shown that algae can be grown in a bioreactor using microbubbles to supply nutrients and keep the cells floating. The microbubbles are produced by a novel technology developed by the team. Not only that, but the same system can be used to supply ozone, which kills bacterial contaminants, and then, by altering the microbubble size, to bring the algae to the surface and therefore make it much simpler to extract the lipids from the algae. This work was published in Algal Research vol 17 pages 217-226.
July 2016 - Prevention of bacterial adherence to human skin
A major problem for some patients, such as the elderly and diabetics, is infection of skin wounds and ulcers by bacteria such as methicillin resistant Staphylococcus aureus, MRSA. An interdisciplinary group including Dr Lynda Partridge in MBB and groups in Medicine and Materials Engineering, has published research in PLOS One July 2016 page e0160387, showing that human proteins called tetraspanins can be used to coat the surface of human skin cells and reduce the adherence of bacteria, allowing them to be washed off. The work was funded by Age UK, and the group are now working towards developing an antibacterial dressing. he picture shows tetraspanins (yellow) coating human skin cells (blue).
June 2016 - How to create a double-stranded DNA copy
It has been known for a long time that when DNA is copied, it can only be copied in one direction, from 5' to 3'. This means that one strand (the 'leading strand') is copied continuously, but the other one (the 'lagging strand') is copied in short sections called Okazaki fragments. These have a short overhang or 'flap' (in pink below left), which then has to be cut off before the fragments can be joined together. This is done by an enzyme called flap endonuclease. A long-term collaboration between Dr John Rafferty in MBB and Prof Jon Sayers in the Medical School at Sheffield has finally succeeded in obtaining an X-ray structure of flap endonuclease in action. It has a remarkable 'arch' which loops out to allow the flap to enter, after which it closes, dragging the flap close to the enzyme active site where the DNA is cut, before the flap opens out again to allow the DNA to leave and prepare for the next round. The work was published in Nature Structural & Molecular Biology 2016, doi:10.1038/nsmb.324, where it made the front cover!
May 2016 - new anticancer drug target
Cancer is not a single disease and invariably requires more than one biochemical lesion to lead to active cancer. There is increasing recognition that a good way to tackle many cancers is to use a cocktail of drugs, each targetting different aspects of the disease. In Scientific Reports vol 6 page 26626, Prof Sherif El-Khamisy and his team have shown that the compound isoeugenol, extracted from the essential oil of a number of plants including ylang-ylang, inhibits a specific route for DNA repair, and could be a novel type of anticancer drug, particularly if used in combination with one of the camptothecin group (camptothecin, irinotecan or topotecan), which target a different but related route.
April 2016 - understanding autosomal polycystic kidney disease
Autosomal polycystic kidney disease produces cysts on the kidney, affects about 1 in 600 people worldwide, and is a major cause of kidney failure. In a collaboration with colleagues in the Medical School in Sheffield, Prof Williamson has investigated a protein linked to the disease (J Am Soc Nephrol 2016 vol 27 issue 4 page 1159). They concentrated on a small part of it that has not been looked at before called the PLAT domain (right), and showed that it binds to lipids in membranes (cyan and green/red molecules), differently depending on phosphorylation (at the amino acid coloured red) and on calcium (the orange ball at the top). This switches the entire protein between being at the cell surface and inside the cell. This discovery opens a new approach to curing the disease.
March 2016 - spatial oscillations in bacteria
E. coli cells are rod-shaped. They grow by getting longer, and when they are long enough, they split in the middle to create two new cells. How does an E. coli cell know where the 'middle' is? It turns out that they have a set of proteins that oscillate regularly from one end to the other (amazingly, roughly once every 25 seconds), and the 'middle' is the place where there is least of these proteins. The key components of this oscillator are called MinD and MinE. A paper published by new lecturer Ling Hwang and colleagues in Proceedings of the National Academy of Sciences of the USA (March 2016, volume 113, issue 11, page E1479) has shown how MinD and MinE work, and has shown that they can be taken out of E. coli and used to create beautiful oscillatory patterns on surfaces. The image shows patterns described by the authors as (left to right) amoebas, waves, spirals, spirals and mushrooms.
February 2016 - nanoscale patterning of proteins on surfaces
We are familiar with the idea of depositing atoms or molecules onto microchips: this is the basis of the modern electronics industry. However, depositing proteins on microchips remains a major technical challenge - one that needs to be overcome to generate (for example) artificial photosynthesis on a chip. Prof Hunter published an elegant method for nanoscale patterning in Feb 2016 (Langmuir volume 32 issue 7 page 1818), which is based on industry-standard light masks and could have wide applications.
January 2016 - how cells decide what size and shape to be
Fundamental questions in biology are: how do cells know what shape to be, when to stop growing and divide, and how to organise themselves? A recent paper, from Prof Simon Foster and Dr Roy Chaudhuri and colleagues, has provided an answer to this question. They studied the spherical bacterium Staphylococcus aureus, a major hospital superbug, and discovered a group of proteins (one of which is the protein PlsY, shown in the image), which distribute themselves in a regular pattern around the outside of the cell. The key result, obtained by combining experimental observations with mathematical modelling, is that the information on where to go arises spontaneously within the cell, as a result of competition between the curvature that these proteins naturally impose on the cell wall, and random thermal motions. Thus, local interactions build up naturally to determine the overall cell shape and provide structural anchors within the cell, even though the proteins involved are much smaller than the overall cell dimensions. The work was published in Proceedings of the National Academy of Sciences of the USA 2015 vol 112 issue 51, page 15725.
December 2015 - photosynthesis in plants
The energy neeeded for almost all life on earth ultimately comes from photosynthesis. The key processes of photosynthesis take place in specialised membrane structures called thylakoids, which not only carry out the capture of light but convert it efficiently to chemical energy, tune the process to environmental conditions such as light intensity, and make sure that the photosynthetic energy does not get diverted into other routes, such as oxidative damage. MBB's Dr Matt Johnson has published a review in the journal Nature Plants (2015, vol 1, article 15161), discussing recent results that demonstrate the dynamic nature of thylakoids. In a second paper, also published in Nature Plants together with MBB professors Neil Hunter and Peter Horton (2015, vol 1, article 15176), he has shown that there is a carefully regulated interplay between the light harvesting apparatus that captures sunlight and the photosystems that convert it to chemical energy, to maximise efficeincy and minimise oxidative damage.