A Visit to Diamond Light Source

Last February, a group of Birkbeck students were given the opportunity to visit the UK's synchrotron light source, Diamond, located near Didcot in Oxfordshire. Most of the students on the trip were taking the Techniques in Structural Biology (TSMB) course as part of a Master's degree. The majority were studying TSMB as a module of the face-to-face M.Sc. in Analytical Bioscience; one was on the distance-learning Structural Molecular Biology M.Sc and a few were studying Bio-Business (which does not include TSMB). This photo shows most of the group with Professor Nick Keep, director of the Structural Molecular Biology course and Dr Katherine Thompson, director of Analytical Bioscience, with part of Diamond's main building in the background.

Birkbeck staff and students in front of the 'doughnut' that houses the Diamond ring system

Dr David Price from Diamond gave the students a short lecture to introduce Diamond and the types of experiment that can be carried out there.

He explained that Diamond is a synchrotron radiation source, which means that it accelerates beams of electrons around the rim of a large doughnut-like structure so they reach speeds approaching that of light and give out extremely intense electromagnetic radiation in the form of X-rays 100 billion times more intense than the sun's rays. X-rays have wavelengths that range from 10^-8 to 10^-11 m, and those in the middle of this range, with wavelengths close to the length of inter-atomic bonds. These are ideally suited to probe the structure of matter on the molecular level, and they can do so in a lot of different ways.

Price explained briefly how a synchrotron radiation source works. Electrons are fired from an 'electron gun' into a linear accelerator and then into a small ring known as the booster synchrotron for further acceleration. They then enter the large storage ring, which has a circumference of about half a kilometre; it is this that gives the overall structure its doughnut-like shape. Strictly speaking, however, it is not a smooth ring but a polygon with 48 straight sections. The electron beam is bent by magnetism at each of its vertices, and it is this process that emits the intense X-rays. The X-rays produced at each vertex are emitted through a 'hole in the wall' at that vertex. They are then filtered and focused using complex optical equipment to give them the exact properties that are needed for a particular experiment. Each of these 'beamlines' is essentially a small laboratory with its own optics and analysis equipment and its own staff team. As the synchrotron structure has 48 vertices, there are 48 possible beamline stations, although they are not all operational at the same time. The Diamond website currently lists 31 operational beamlines, of which seven have been optimised for macromolecular X-ray crystallography. Some of the other techniques available are absorption and fluorescence spectroscopy and small-angle X-ray scattering.

There are more than 50 synchrotron radiation sources ('lightsources') in the world; Diamond is one of the more powerful and versatile of these, but it is the only one in the UK. Most - about 90% - of its users are academics who are required to publish their work in open access journals; over 1000 publications each year cite Diamond Academics apply for time on a specific beamline, and these applications are peer reviewed. About 10% of users come from industry and pay for access.

Price also highlighted briefly a few recent examples of research carried out at Diamond, focusing mainly but not exclusively on structural biology. These included the structure of a peptide hormone, GLP-1 (PDB 5NX2), that stimulates the secretion of insulin by beta cells in the pancreas and that could is an important target for drugs to treat type 2 diabetes. The structure of the EV71 virus (PDB 4CDQ), which causes the potentially fatal hand, foot and mouth disease, was solved at Diamond in 2012 and this is now being used to design potential drugs for this occasionally fatal disease. Away from structural biology - but not away from medicine - he described structural studies of metal alloys that can help understand why hip replacements can be rejected and inform the design of improved materials.

After the talk, the Birkbeck party was divided into two smaller groups to tour of the main site. Both groups visited one of the beamlines devoted to macromolecular X-ray crystallography, I24, where the structure of GLP-1 was solved. This beamline includes facilities for determining structures of viruses and membrane proteins, and for working with very small crystals (down to about 1.5 microns in diameter).

Looking down on a small part of the storage ring

Birkbeck staff (Prof. Nick Keep, far left and Dr. Clare Sansom, next left) and students in the control cabin of beamline I24, one of those used for macromolecular X-ray crystallography

The trip ended with a short visit to the UK's high-energy neutron and muon source, located on the same campus as Diamond and named ISIS after Oxford's river. These beams are also used to probe the structure of matter, using techniques such as neutron diffraction. We would like to thank all Diamond and ISIS staff involved for their contributions to a fascinating day.

PPS students will learn more about X-ray diffraction in the second-year module Techniques in Structural Molecular Biology (TSMB), and much more still if they choose to take the specialist Protein Crystallography course. And students on either course may get a chance to visit Diamond fir themselves, as we hope to run the trip again next year.

Welcome to new PPS students - and blogging crystallography

This post is very like those I have written at the beginning of the academic year for the past few years, if posted rather later than usual. This is because what I have to say now is also very similar...

I would like to offer a warm welcome to the Principles of Protein Structure blog to all students who have recently started studying Birkbeck's Principles of Protein Structure (PPS) course, and a welcome back to any who have taken a break in studies and intend to complete the course this year.

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The programme for Autumn 2017 focuses on the molecular biology of cancer; there is some material on this topic in section 5 of this course, 'Towards Tertiary Structure', where we look briefly at the structure and function of kinases. Many of the newer anti-cancer drugs, including Glivec, which has transformed the prospects for patients with chronic myeloid leukaemia, target this class of protein. Other posts may be reports from conferences or summaries of recently published papers in protein structure and allied areas; watch out for one at the end of this year featuring a lecture by the UK's newest Nobel laureate, Richard Henderson.

Some earlier posts were written by "guest blogger" Jill Faircloth, who took the MSc in Structural Molecular Biology a few years ago and is now working as a freelance science communicator. She introduces herself in this post written in March 2012, in which she also describes how she found the later part of the PPS course and her thoughts on the two choices available for the second year of the MSc.

Do, if you get a chance, look through some blog posts from earlier years to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

And I hope that you will also be interested in some more blogging of mine, in which I explore crystallography - the most popular experimental technique for determining protein structures - more widely. In August I was lucky enough to attend the congress of the International Union of Crystallography in Hyderabad, India, and to write the conference blog. The topics I covered over the 8 days of the meeting ranged from crystallography in history to crystallography in space, but did also include some structural biology. The first post in this series covers the opening ceremony, including the award of one of crystallography's highest honours, the Ewald Prize, to Sir Tom Blundell, a former head of the Department of Crystallography at Birkbeck (now part of the Department of Biological Sciences). Sir Tom is perhaps best known for his part in solving the structure of HIV protease, target of some of the most successful drugs for AIDS, and he went on to found a drug discovery company, Astex. A later post describes a plenary lecture by John Spence of Arizona State University on imaging proteins in motion.

Finally, the best of luck for the 2017-18 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom
Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure

The 2017 ISMB Retreat

The Institute of Structural and Molecular Biology (ISMB) at Birkbeck and UCL holds a conference each summer where all members of the Institute's constituent departments come together to discuss their research. In even years, this takes the form of the ISMB Symposium: two intense days of research talks from ISMB core members and invited research leaders. In odd years, the focus turns to the younger members of the Institute with the ISMB Retreat. This, usually held in Cambridge University's Robinson College, is a two-day residential event featuring plenaries from outside speakers, shorter talks by students and postdocs from the ISMB, an activity designed to get young researchers thinking about science-based careers outside an academic lab, and a poster session. The 2017 retreat, held on June 29 and 30, was the seventh in the series.

There is a full report of this retreat (written by me) with photos on the ISMB website. Do check it out! Here are just a few highlights that are relevant to students of structural biology.

The three keynote speakers were Lori Passmore (MRC Laboratory of Molecular Biology (LMB), Cambridge; Bill Rutherford (Imperial College, London); and Bart Vanhaesebroeck (UCL Cancer Institute). Structural biology was a feature of all three talks, although most prominently in Passmore's. She was recently appointed as a group leader at the LMB, where her group uses high resolution electron microscopy to study the process of poly-adenylation (link to PPS section 8): the addition of the 'poly-A tail' to newly synthesised messenger RNA molecules during the maturation process. Much of the work she discussed has still to be published, apart from a section on techniques development: a physicist in her group, Chris Russo, has worked with her to devise a novel, gold-based substrate for mounting specimens in the electron microscope that eliminates most specimen movement and thus increases resolution.

Bill Rutherford gave an engaging talk, unusually featuring hand-written slides, on the mechanism of action of photosystem II, one of the complex proteins involved in photosynthesis. The evolution of photosynthesis was responsible for the increase in oxygen in the Earth's atmosphere that made multicellular life possible, and Rutherford explained how the increase in oxygen had led to further changes in the structure and mechanism of the photosystem that had the overall effect of decreasing its efficiency. Bart Vanhaesebroeck's lecture, which ended the retreat, described the mechanism of the phosphoinositide-3-kinase family of proteins and their role in cancer. Inhibitors of these kinases (link to PPS section 5) might prove useful anti-cancer drugs, almost certainly as part of combination therapy.

The programme also included nine short talks by students and young postdocs. The excellent quality of these was highlighted by the judges of the best talk, who were unable to come to a consensus judgement. In the end, two equal prizes were given, to Jennifer Booker, who studies the structure of sodium channels (see e.g. this post from September 2016) in Bonnie Wallace's group at Birkbeck, and to Sapir Ofer, for a talk on her PhD studies of the structural and molecular biology of archaeal histones. Proteins in this family are responsible for packing DNA into the chromosomes of eukaryotic cells; bacteria pack DNA using an entirely different mechanism, but the mode of packing in archaea - single-celled organisms that have no nucleus and are defined as prokaryotes but are more closely related to eukaryotes than to bacteria - was unknown until recently.

The retreat always includes an activity to challenge ISMB students and postdocs to think about career opportunities beyond academic research, and this year's proved particularly popular. It was a Dragons' Den style competition in which the younger delegates were split into teams, assigned mentors and given an hour to create a fictitious life science company, develop a bid for funding and present this to a panel of 'dragons'. Three finalists were chosen to battle it out in front of the rest of the delegates. The competitors worked hard and all 'companies' produced ideas that held water to at least some extent, but there could be only one winner. That was a company named TerraNova, which presented an antibody-drug conjugate for primary progressive multiple sclerosis.

Highlights from the summer 2017 ISMB seminar programme

Regular readers of this blog will know that the Institute of Structural and Molecular Biology (ISMB) coordinates the research efforts in the Department of Biological Sciences at Birkbeck and two departments – Chemistry and Structural and Molecular Biology – at neighbouring University College London. Research in the participating departments is coordinated through six core research centres, and many grants, PhD studentships and experimental facilities are held in common.

Since 2010, too, these departments’ seminar programmes have been consolidated into termly series of ISMB seminars, giving the Institute’s researchers and students the chance to hear world-class scientists present their work. Most of each term’s seminars are centred round a theme, with recent themes including bioinformatics; the molecular basis of infectious disease; and, in the most recent series, ‘beyond signalling’. This post briefly describes two seminars in this series, both from researchers based in London and both closely concerned with disease mechanisms.

Paul Freemont holds a chair in protein crystallography at Imperial College, London. His group has solved the structure of several proteins linked to cancer, including a domain called the ‘RING finger’ that is found in the breast cancer susceptibility protein BRCA1. His talk to the ISMB, however, concerned a research interest that he shares with the Institute’s head, Gabriel Waksman: the membrane-bound protein complexes through which Gram negative bacteria secrete toxins and other molecules across the double bacterial cell wall and out of the cell. These bacteria have evolved at least six such ‘molecular machines’, with yet another found in mycobacteria such as M. tuberculosis. Waksman’s work in elucidating the structure of the Type IV system is covered extensively in section 11 of the PPS course (‘Structures of Membrane Proteins’).

Freemont’s seminar described the Type VI secretion system (T6SS), which was first identified as recently as 2006 in Vibrio cholerae (as its name suggests, the causative agent of cholera). The function of the T6SS is to eject proteins from the interior of the bacterial cell into an adjacent cell, which may be either bacterial or eukaryotic. Freemont’s lab mainly studies these systems in the bacterium Pseudomonas aeruginosa, an opportunistic pathogen that causes infections mainly in people who are already chronically ill, such as cystic fibrosis patients and those with severe burns.

This secretion system has been described as a ‘molecular syringe’. Its structure resembles that of the tail of a bacteriophage – a type of virus that affects bacteria – but it is inverted, with the tip of the tail pointing away from the bacterial cell wall and towards its target cells. In some species, the same secretion systems can target both eukaryotic cells and other bacteria. The system consists of a long sheathed tube, built up from many protein subunits, that is large enough to be easily viewed using electron tomography and that is tipped by a spike through which the protein to be delivered is ejected. Energy for cargo delivery is provided by the contraction of the tube, with a single contraction storing the energy equivalent of 1600 molecules of ATP. The whole structure is dynamic; it is assembled only when needed and disassembled after the cargo has been delivered, allowing the cycle to begin again.

Although the T6SS can sometimes act as a cell-to-cell ‘killing machine’, as in Vibrio cholerae, protein delivery to the target cell will often have rather more subtle effects, with Pseudomonas aeruginosa a case in point. This rod-shaped Gram negative pathogen uses three distinctly different type 6 systems, encoded on separate operons, to secrete effector proteins that interfere with the host immune system. Freemont and his group have solved the structures of several P. aeruginosa T6SS components using X-ray crystallography, throwing more light on their phage-like mechanism of action. Structures of an accessory protein (TagJ) and the ATPase that catalyses sheath disassembly (ClpV) were all published in the Journal of Biological Chemistry in 2014; some other component structures are yet to be published. TagJ is now known to interact both with ClpV, an AAA+ ATPase, and with components of the sheath, and this interaction allows the rapid disassembly that is required for the complete system to be reset. Each ATPase only interacts with the components from its own operon. Further structural studies, using high-resolution electron microscopy as well as X-ray crystallography, are expected to elucidate further details of these complex molecular machines and to suggest ways in which they might one day be targeted by the novel antimicrobial drugs that we so desperately need.

The second London-based seminar speaker, Miriam Dwek from the University of Westminster, had a somewhat unorthodox beginning to her research career at Oxford University’s first spin-off company, Oxford Glycosystems (now, after many mergers, part of pharma giant UCB). She has maintained her interests in glycobiology (the biology and biochemistry of sugars and polysaccharides) and its application to human disease – particularly breast cancer – into and throughout her academic research career.

Breast cancer is one of the most common cancer types, with 400,000 new cases occurring each year in Europe alone. Breast tumours can be divided into many subtypes with different genetic and biochemical profiles; although some are now easily treated with surgery, radiotherapy and/or drugs, others are often fatal (if perhaps after many years). Generally speaking, tumours are tractable when they are confined to breast tissue and the disease only becomes difficult to treat once it has spread. All cancer types metastasise in a particular pattern; breast cancers tend to spread first into nearby lymph nodes and then to the lungs, brain or bones.

Metastasis is a complex, multi-step process, and selecting the optimum treatment for each patient depends on detecting whether and how her tumour will metastasise as early as possible. Changes in the concentration of some biological molecules in body fluids have been associated with tumour growth and development, and these biomarkers can be used as easily-measurable surrogates of cancer development. One particularly well-known example is the prostate-specific antigen (PSA), a glycoprotein found in semen that is elevated in prostate cancer. No such clear-cut examples exist in breast cancer, but many subtler biochemical changes are known to occur. Dwek and her group have been exploring differences in protein glycosylation patterns between breast tumours and normal breast cells.

Glycosylation is one of the most common post-translational modifications of amino acids (link is to PPS section 2). There are two basic types; one or (almost always) more monosaccharides can be bonded to the oxygen atoms of serine and threonine side chains (O-glycosylation) or to asparagine’s side chain nitrogen (N-glycosylation). The addition of the first residue to the amino acid and the subsequent extension of the chain are catalysed by enzymes in the transferase class. In O-glycosylation, in particular, the patterns of residues added to the glycan ‘branches’ differ between healthy breast epithelial cells and breast tumour cells, and this, in turn, can aid the process of cell adhesion (binding cells together), which is essential for tumour metastasis. Cadherins are glycoproteins that have important roles in cell adhesion, and Dwek’s group used glycoproteomics techniques to identify this as a potential biomarker of metastatic breast cancer. She considers that it is likely to be particularly useful for detecting metastasis in patients with estrogen receptor positive tumours and vascular invasion.

Other topics presented by leading researchers in this ISMB seminar series included nuclear receptors, collagen-binding proteins and protein targeting and translocation. The seminar programme will return in October, and I will doubtless be returning to it again in this blog.

Welcome to PPS students 2016-17!

This post is extremely like those I have written at this time of year for the past few years. This is because what I have to say now is very, very similar...

I would like to offer a warm welcome to the Principles of Protein Structure blog to all students who have just started studying Birkbeck's Principles of Protein Structure (PPS) course, and a welcome back to any who have taken a break in studies and intend to complete the course this year.

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The programme for Autumn 2016 focuses on the molecular and structural biology of infectious disease; there is material on similar topics in section 10 of the PPS course, 'Protein Interactions and Function'. Other posts may be reports from conferences or summaries of recently published papers in protein structure, protein bioinformatics and allied areas. Look out for an account of a conference on structural assemblies at Birkbeck in December that will honour the 50-year career of one of our emeritus professors, Steve Wood.

Some earlier posts on this blog were written by "guest blogger" Jill Faircloth, who took the MSc in Structural Molecular Biology a few years ago and is now working as a freelance science communicator. She introduces herself in this post written in March 2012, in which she also describes how she found the later part of the PPS course and her thoughts on the two choices available for the second year of the MSc.

Do, if you get a chance, look through some blog posts from earlier years to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

I particularly recommend that you look at a couple of posts from December 2013 and July 2014 about the history of structural science, particularly X-ray crystallography. Crystallography was the first method to be developed for solving the structure of biological macromolecules, and it is still the most important. The year 2014 was designated by the United Nations as the International Year of Crystallography, marking the year between the centenaries of the publication of the first papers on X-ray diffraction and the award of the 1915 Nobel Prize for Physics to the father-and-son team of William and Lawrence Bragg who made the principal discoveries.

So - the best of luck for the 2016-17 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom
Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure

Welcome to PPS students 2015-16!

This post is extremely like those I have written at this time of year for the past few years. This is because what I have to say now is very, very similar...

I would like to offer a warm welcome to the Principles of Protein Structure blog to all students who have just started studying Birkbeck's Principles of Protein Structure (PPS) course, and a welcome back to any who have taken a break in studies and intend to complete the course this year.

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The programme for Autumn 2015 focuses on transcription, which is the process through which DNA is copied into RNA; this will be covered in some detail in one of the later sections of the course, the Protein Lifecycle. Other posts may be reports from conferences or summaries of recently published papers in protein structure, protein bioinformatics and allied areas. Look out for an account of a lecture to be given at Birkbeck in October by Venki Ramakrishnan, who was awarded a share of the Nobel Prize for Chemistry in 2009 for structural studies of ribosomes and will shortly take over as President of the Royal Society.

Some posts on this blog are written by "guest blogger" Jill Faircloth, who took the MSc in Structural Molecular Biology a few years ago and is now working as a freelance science communicator. She introduces herself in this post written in March 2012, in which she also describes how she found the later part of the PPS course and her thoughts on the two choices available for the second year of the MSc.

Do, if you get a chance, look through some blog posts from earlier years to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

I particularly recommend that you look at a couple of posts from December 2013 and July 2014 about the history of structural science, particularly X-ray crystallography. Crystallography was the first method to be developed for solving the structure of biological macromolecules, and it is still the most important. The year 2014 was designated by the United Nations as the International Year of Crystallography, marking the year between the centenaries of the publication of the first papers on X-ray diffraction and the award of the 1915 Nobel Prize for Physics to the father-and-son team of William and Lawrence Bragg who made the principal discoveries.

So - the best of luck for the 2015-16 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure

Microtubules and Microscopes: Exploring the Cytoskeleton

Electron microscopist Carolyn Moores, the most recently appointed professor in the Department of Biological Sciences at Birkbeck, gave her inaugural lecture at the college on June 1.

Moores arrived at Birkbeck in 2004 to start her research group and has risen rapidly and steadily up the academic ladder ever since. Introducing the lecture, the Master of Birkbeck, David Latchman, explained that Moores’ CV stood out in every way; she was clearly as gifted a teacher and administrator as she was a researcher. Furthermore, as she has won several awards for science communication, he predicted that the audience would be in for a treat. We were not disappointed.

Moores began her lecture by saying that she would talk about three different things: her own career development; her group’s research into the structure and function of microtubules; and the advancement of women in science, a cause that is close to her heart.

She remembered that she had wanted to work as a scientist as soon as she knew what a laboratory was, and she started young, as an intern in a research lab at Middlesex Hospital while still in the sixth form. School was followed by a BSc in Biochemistry at Oxford and a PhD in John Kendrick-Jones’ lab at the world-famous Laboratory for Molecular Biology (LMB) in Cambridge. She then moved to work as a post-doc with Ron Milligan at the Scripps Research Institute in La Jolla, California, USA, and it was there that she began her studies on microtubules.

The award of a David Phillips research fellowship in 2004 gave her the opportunity to return to the UK as an independent researcher. She explained that there were three reasons – or more accurately three people – that led her to choose to come to Birkbeck. Working in electron microscopy, she was inspired by the work of Helen Saibil, one of the UK’s principal exponents of that technique; she had known Nicholas Keep, then a lecturer in Biological Sciences, as a friend since her time at the LMB; and she knew that she would value the interdisciplinary working environment of the Institute for Structural Molecular Biology under the ‘inspired’ leadership of Gabriel Waksman.

Moores then moved on to discuss the main topic of her group’s research: the three-dimensional structure, function and role in disease of tiny cylindrical structures known as microtubules. These are one of the building blocks of the cytoskeleton, which forms a framework for our cells in the same way that our skeletons form a framework for our bodies. They are about 25nm in diameter, which puts them firmly into the ‘nano-scale’ of biology that is easily studied using electron microscopy.

There is a cytoskeleton in every living cell, and it, and the microtubules that form it, are involved in many important cellular processes including shape definition, movement and cell division. Diseases as diverse as cancer, epilepsy, neurodegeneration and kidney disease have been linked to microtubule defects. Understanding their fundamental structure and function, as Moores’ group aims to, should help in understanding these disease processes and perhaps also in developing effective treatments.

Microtubules are built up from many copies of a small protein called tubulin, which, in turn, is a dimer of two similar proteins called alpha and beta tubulin. These tubulin dimers make contacts with each other both head-to-tail and side-to-side to create the cylindrical microtubule wall, fuelled by energy derived from the molecule GTP. Each tubulin unit has a definite “top” and a “bottom” and, as the units are oriented in parallel, so has the complete microtubule.

Microtubules are dynamic structures; they continue to grow by the addition of tubulin units to one end as long as GTP is available, and then begin to unravel and shrink. This dynamism, which allows them to respond to the changing needs of the cell, is essential for their function in healthy cells. In particular, microtubules organise chromosome structures during cell division and are therefore necessary for cell proliferation. As cancer is a disease of uncontrolled cell proliferation, it is possible to imagine that a molecule that could specifically block microtubule growth and assembly in the nucleus might be useful as an anti-cancer drug.

Moores and her group are aiming to understand the process of microtubule growth at as high resolution as possible, using electron microscopy. Unfortunately, however, the most detailed images can only be obtained if the specimen is at very low temperatures (in so-called cryo-elecron microscopy) and using this means that the dynamics of the specimens must be “frozen” into a still image. While it is now possible to see the individual tubulin subunits in the static microtubule images, many details of their structure can only be inferred from computational analysis.

Moores went on to describe one project in her lab in a little more detail. This was an investigation of the structure and role of proteins that bind only to growing microtubule ends, falling off when the growth stops. It is possible to obtain low-resolution images of microtubules in which these molecules have been made to fluoresce, so only growing microtubules are tracked.

In order to understand the growth process in detail, the group developed an analogue of the GTP “fuel” molecule which can bind to the tip of a microtubule that is extending but not break down to release its energy, so the microtubule does not in fact grow. This forms a static analogue of a growing microtubule that retains all the characteristics of the dynamic structure but that can be studied at low temperatures.

The binding site for End Binding protein 1 (highlighted in green) on the microtubule lattice at the corner of four tubulin dimers, visualised using cryo-electron microscopy
Image from Maurer et al. 2012, Cell 149(2):371-82. Full text here.

Images of this structure have shown that the end binding proteins bind at the corner of four of the tubulin units. They have explained a lot of the properties of growing microtubules, but there is still more to learn. A full understanding will need structures that are at even higher resolution, where the positions of individual atoms can be made out. Following many years of technical development, today’s most powerful electron microscopes are now making this possible.

In the last section of the lecture, Moores left the topic of research to talk briefly about another of her passions: the promotion of women in science. She explained that although 65% of under-graduates in the biological sciences are now women, the proportion of women drops to 40% at any academic grade and 25% for full professors.

A study cited by the European Molecular Biology Organisation has suggested that the barriers for women scientists to progress are set so high that at the current rate of progress full equality would never be achieved. Birkbeck has signed up to the Athena SWAN Charter, set up to encourage higher education institutions to transform their culture and promote gender equality. She described her work with the Athena SWAN team that has so far resulted in the college gaining a bronze award as being as exciting as, but also as challenging as, her studies of microtubules.

There is more information about the structure and function of microtubules on this page in section 7 of the PPS course. The technique of cryo-electron microscopy is covered in some detail in the second-year MSc module Techniques in Structural Molecular Biology.

Welcome to PPS students 2014-15!

This post is extremely like those I have written at this time of year for the past few years. This is because what I have to say now is very, very similar...

I would like to offer a warm welcome to the Principles of Protein Structure blog to all students who have just started studying Birkbeck's Principles of Protein Structure course, and a welcome back to any who have taken a break in studies and intend to complete the course this year.

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. Throughout the taught course (but more often in the later part of the course) I will post reports of recent developments. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The overall title of the programme for Autumn 2014 is Synthetic Biology: an innovative and important topic that relates quite closely to some of the material we cover in the later sections of the course, particularly the sections on Bioinformatics and the Protein Lifecycle. Other posts may be reports from conferences or summaries of recently published papers in protein structure, protein bioinformatics and allied areas.

Some of the posts on this blog are written by "guest blogger" Jill Faircloth, who took the MSc in Structural Molecular Biology a few years ago and is now working as a freelance science communicator. She introduces herself in this post written in March 2012, in which she also describes how she found the later part of the PPS course and her thoughts on the two choices available for the second year of the MSc.

Do, if you get a chance, look through some of the earlier blog posts to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

I particularly recommend that you look at a couple of posts from last academic year - December 2013 and July 2014 - about the history of structural science, particularly X-ray crystallography. Crystallography was the first method to be developed for solving the structure of biological macromolecules, and it is still the most important. The year 2014 was designated by the United Nations as the International Year of Crystallography, marking the year between the centenaries of the publication of the first papers on X-ray diffraction and the award of the 1915 Nobel Prize for Physics to the father-and-son team of William and Lawrence Bragg who made the principal discoveries. The International Year has been marked by a wide range of activities, special symposia, publications, "open labs" and even postage stamps, and there are still a few events planned.

So - the best of luck for the 2014-15 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure

Science Week 2014: Birkbeck and the History of Crystallography

Science Week at Birkbeck in 2014 featured two lectures on Department of Biological Sciences, both presented on 2 July. One of these was a double act from two distinguished emeritus professors and Fellows of the College, Paul Barnes and David Moss. Remarkably, they both started their working lives at Birkbeck on the same day – 1 October 1968 – and so had clocked up over 90 years of service to the college between them by Science Week 2014.

The topic they took was a timely one: the history of the science of crystallography over the past 100 years. UNESCO has declared 2014 to be the International Year of Crystallography in recognition of the seminal discoveries that started the discipline, which were made almost exactly 100 years ago; a number of the most important discoveries of that century were made by scientists with links to Birkbeck.

The presenters divided the “century of crystallography” into two, with Barnes speaking first and covering the first 50 years. In giving his talk the title “A History of Modern Crystallography”, however, he recognised that crystals have been observed, admired and studied for many centuries. What changed at the beginning of the last century was the discovery of X-ray diffraction. Wilhelm Röntgen was awarded the first-ever Nobel Prize for Physics for his discovery of X-rays in 1896, but it was almost two decades before anyone thought of directing them at crystals. The breakthroughs came when Max von Laue showed that a beam of X-rays can be diffracted by a crystal to yield a pattern of spots, and the father-and-son team of William Henry and William Lawrence Bragg showed that it was possible to derive information about the atomic structure of crystals from their diffraction patterns. These discoveries also solved – to some extent – the debate about whether X-rays were particles or waves, as only waves diffract; we now know that all electromagnetic radiation, including X-rays, can be thought of as both particles and waves

Von Laue and the Braggs were awarded Nobel Prizes for Physics in 1914 and 1915 respectively, and between 1916 and 1964 no fewer than 13 more Nobel Prizes were awarded to 18 more scientists for discoveries related to crystallography. Petrus Debye, who won the Chemistry prize in 1936, showed how to quantify the thermal motion of atoms as vibrations within a crystal. He also invented one of the first powder diffraction cameras, used to obtain diffraction patterns from powders of tiny crystallites. Another Nobel Laureate, Percy Bridgman, studied the structures of materials under pressure: it has been said that he would “squeeze anything he could lay his hands on”, often up to intense pressures.

Scientists and scientific commentators often argue about which of their colleagues would have most deserved to win the ultimate accolade. Barnes named three who, he said, could easily have been Nobel Laureates in the field of crystallography. One, Paul Ewald, was a theoretical physicist who had studied for his PhD under von Laue in Munich, and the other two had strong links with Birkbeck. JD “Sage” Bernal was Professor of Physics and then of Crystallography here; he was famous for obtaining, with Dorothy Crowfoot (later Hodgkin) the first diffraction pattern from a protein crystal, but his insights into the atomic basis of the very different properties of carbon as diamond and as graphite were perhaps even more remarkable. He took on Rosalind Franklin, whose diffraction patterns of DNA had led Watson and Crick to deduce its double helical structure, after she left King’s College, and she did pioneering work on virus structure here until her premature death in 1958.

Barnes ended his talk and led into Moss’s second half-century with a discussion of similarities between the earliest crystallography and today, as now, you only need three things to obtain a diffraction pattern: a source of X-rays, a crystalline sample, and a recording device; the differences all lie in the power and precision of the equipment used. He demonstrated this with a “symbolic demo” that ended when he pulled a model structure of a zeolite out of a large cardboard box.

Paul Barnes demonstrates the basic principles of X-ray crystallography using a large cardboard box. Photo © Harish Patel and Ruben Zamora, Department of Psychological Sciences, Birkbeck

David Moss then took over to describe some of the most important crystallographic discoveries from the last half-century. His talk concentrated on the structures of large biological molecules, particularly proteins, and he began by explaining the importance of protein structure. All the chemistry that is necessary for life is controlled by proteins, and knowing the structure of proteins enables us to understand, and potentially also to modify, how they work.

Even the smallest proteins contain thousands of atoms; in order to determine the position of all the atoms in a protein using crystallography you need to make an enormous number of measurements of the positions and intensities of X-ray spots. The process of solving the structure of a protein is no different from that of solving a small molecule crystal structure, but it is more complex and takes much more time. Very briefly, it involves crystallising the protein; shining an intense beam of X-rays on the resulting crystals to produce diffraction patterns, and then doing some extremely complex calculations. The first protein structures, obtained without the benefit of automation and modern computers, took many years and sometimes even decades.

Thanks to Bernal’s genius, energy and pioneering spirit, Birkbeck was one of the first institutes in the UK to have all the equipment that was needed for crystallography. This included some of the country’s first “large” computers. One of the first electronic stored-program computers was developed in Donald Booth’s laboratory here in the 1950s. In the mid-1960s the college had an ATLAS computer with a total memory of 96 kB. It occupied the basements of two houses in Gordon Square, and crystallographers used it to calculate electron density maps of small molecules. Protein crystallography only “took off” in the 1970s with further improvements in computing and automation of much of the experimental technique.

Today, protein crystallography can almost be said to be routine. The first step, crystallising the protein, can still be an important bottleneck, but data collection at powerful synchrotron X-ray sources is extremely rapid and structures can be solved quite easily with user-friendly software that runs on ordinary laptops. There are now over 100,000 protein structures freely available in the Protein Data Bank (PDB), and about 90% of these were obtained using X-ray crystallography. The techniques used to obtain the other 10,000 or so, nuclear magnetic resonance and electron microscopy, are more specialised.

Moss ended his talk by describing one of the proteins solved in his group during his long career at Birkbeck: a bacterial toxin that is responsible for the disease gas gangrene (PDB 1CA1). This destroys muscle cells by punching holes in their membranes, and its victims usually have to have limbs amputated to save their lives. Knowing the structure has allowed scientists to understand how this toxin works, which is the first step towards developing drugs to stop it. But you can learn even more about how proteins work if you also understand how they move. Observing and modelling protein motion in “real time” still poses many challenges for scientists as the second century of crystallography begins.

Structure of alpha-toxin, the key Clostridium perfringen toxin in gas gangrene. Image from the PDB.

Crystallins under the Lens

Written by Jill Faircloth

For generations, anyone who argued against evolutionary theory would point to the human eye and exclaim that nothing so perfectly adapted to its purpose could have evolved in a series of random steps. The well-rehearsed counter argument is that even a very basic recognition of light and shadow via an organic pinhole camera is useful as an aid to survival and that this could provide the first stepping stone towards the sophistication of the vertebrate eye (see references 1 and 2). The theory is supported by a succession of organisms with gradually increasing vision.

On a molecular level, the proof is harder to achieve but Christine Slingsby of Birkbeck's Department of Biological Sciences has used crystallography to do just that. In investigating the structure of the proteins of the vertebrate eye lens, Slingsby has not only greatly increased our understanding of their characteristics and mechanisms but also provided fascinating insights into their evolution.

Professor Slingsby's work is featured in several pages in the PPS course: Greek Key Motif, Beta Sandwiches, Lens Proteins and Cataract and Eukaryotic Genomes. Last year she published a paper (reference 3 here) which summarised the key conclusions of her research during the last ten years. This review is available in the Birkbeck e-library here.

Vertebrate lenses comprise layers of highly elongated fibre cells which give transparency and focus but the refractive power is given by high concentrations of transparent proteins from two superfamilies: the alpha crystallins and the beta-gamma crystallins. These proteins, which are all mainly made up of beta strands, have been co-opted from their original functions to generate a functioning lens.

You don’t need to look far to find the probable origin of α crystallins. They are small heat shock proteins (sHsps), molecular chaperones that are present in most types of cell in most organisms. They are upregulated: that is, produced in greater quantities, by cells under environmental stress as part of the protein homeostasis response.

Despite their name, βγ crystallins are unrelated to α crystallins; all crystallins interact to form a refractive index gradient which can vary as required. Apart from the vertebrate lens where they are very prevalent, and in stark contrast to α crystallins, βγ crystallins are found only in other vertebrate eye tissues (except as a component of a much larger gene/protein known as Aim1) and this makes their origin harder to identify. Beta and gamma crystallins each contain four Greek key motifs organised as two βγ-crystallin domains.

There are several requirements for an eye lens protein. It must be expressed at very high levels, unlike sHsps, so the sHsp gene promoters would have required modification. The proteins must pack tightly and uniformly enough so that there is no irregularity on the scale of the wavelength of light and they must be soluble but must not crystallise or separate into different phases. In addition, lens fibre cells have lost their organelles, which could cause light scattering, and so have no mechanisms for protein repair or disintegration. Accordingly, these proteins need to have a lifespan as long as the vertebrate using them.

One of the main reasons for crystallins having been adopted as lens proteins could be that the two α crystallins are able to dynamically form polymers with highly diverse size and shape. This ability was demonstrated as the first crystal structure of a sHsp revealed a hollow octahedral structure of 24 α crystallin monomers. The next one to be solved showed point group 32 symmetry and was constructed from six dimers arranged in two interlocking rings.

This figure shows the beta-sandwich structure of the alpha-crystallin domain of a monomer, the formation of the dimer with the B6 beta strand exchanging into the partner beta-sandwich, and the oligomer with six dimers forming interconnecting discs. The dimers link using motifs on the C terminal extension which insert into the pocket between the B4 and B8 strands, shown in dark blue, and by interaction of the N terminal helices.

Figure taken from Slingsby, C. et al. (2013. PDB 1GME

In addition to the wide range of alpha-crystallin oligomers, the numerous βγ-crystallin chains can be assembled to create a wide range of polymers which coexist in a polydisperse stable but flexible arrangement of varying density.

Beta-crystallins thus appear to function in a similar way to α-crystallins, forming a diverse range of differently sized hetero-oligomers that adjust the refractive index throughout the lens.

Gamma crystallins are different because they are monomeric and polar. They are present in differing concentrations throughout the lens and their polarity results in distinctive orientations towards other crystallins which may regulate inter-crystallin interactions. There is evidence that disruption of these dipoles results in cataracts.

By examining genomes of organisms which predate the development of the camera eye, Slingsby has shed light on the evolutionary pathway of crystallins as lens proteins. PPS students will bave read about the single-domain βγ-crystallin in the urochordate (invertebrate) sea squirt, Ciona intestinalis, that has exactly the same double Greek key structure as a vertebrate crystallin but includes a calcium binding sequence in each Greek key motif. This ancestral link was further demonstrated by the remarkable discovery that the gene promoter for Ciona-crystallin could successfully target reporter gene expression for proteins associated with vision in vertebrates.

Investigation of the genome of a cephalochordate, which is part of the lineage of both vertebrates and urochordates, revealed a less complex ancestor to βγ-crystallins. Signature sequences from the βγ-crystallins have also been found in bacterial and archaeal proteins. The implication of this is that all of the proteins of the vertebrate lens could well have evolved from proteins present in ancient species with no visual function. An interesting twist is introduced by the knowledge that the nonchordates, or animals without a spine, can use quite similar cellular lenses that involve non-crystallin proteins.

This suggests that lenses evolved independently in different animal kingdoms, relatively late on an evolutionary timescale, utilizing different proteins that were available in the respective phyla, that is proteins which already had an established purpose but which had qualities allowing them to form lenses. Since all species seem to have had access to at least a basic form of βγ-crystallin, it is an impressive demonstration that evolution can not only capitalise on the multiple possibilities presented by one family of proteins to develop a functioning visual system, but also repeat the trick from a different starting point.

References

1. Dawkins, R. (1994). The eye in a twinkling. Nature 368, 690-691
2. Nilsson, D.E., Pelger, S. (1994). A pessimistic estimate of the time required for an eye to evolve. Proc. Biol. Sci. 256(1345): 53-8.
3. Slingsby, C., Wistow, G.J. and Clark, A.R. (2013). Evolution of crystallins for a role in the vertebrate eye lens. Protein Sci. 22(4):367-80.

A Very Short History of Crystallography

You might possibly have been intrigued to read in my last post that 2014 has been designated as the International Year of Crystallography. This year was chosen to celebrate the fact that this discipline - the study of atomic and molecular structure through their crystal forms - is now almost exactly a hundred years old. Admittedly, the first paper in the discipline, rather charmingly titled just "The diffraction of short electro-magnetic waves by a crystal" was published in 1913, and the Nobel Prize awarded two years later, but 2014 is at least a good compromise.

It can be said, perhaps simplistically, that crystallography was invented by the father-and-son team of William Henry and William Lawrence (known as Lawrence) Bragg, at the Universities of Leeds and Cambridge in the UK. The Braggs, however, did not aim to found a new discipline or even to investigate the atomic properties of matter. They were more interested in solving a problem that had been puzzling the cleverest physicists in the world for almost two decades. X-rays had been discovered by Wilhelm Rőntgen in Germany in 1895, but their very name (the unknown X) suggests reveals their controversial nature. Were they particles or waves?

The older Bragg, William, was convinced that X-rays were particles, and set out to prove this to his son (who favoured the wave theory) by exploiting the discovery of another German physicist, von Laue, that X-rays shone at a crystal were scattered and could produce a pattern on a film. Lawrence was the first to realise that these patterns could be explained by the theory that the X-rays were reflected from planes of atoms in the crystal and interfered with each other.

Lawrence presented these results to the Cambridge Philosophical Society late in 1912 and published them in the paper mentioned above the following year. This paper also included the first formulation of one of the best known of all laws of physics: Bragg's Law. This relates the wavelength of incoming X-rays and the angles that they are scattered (diffracted) to the spacing between planes of atoms in a crystal, enabling scientists to determine the geometry of atomic crystal lattices.

The Braggs worked together in Leeds and published their first structures, including that of sodium chloride (common salt) before Lawrence was sent to France to fight in the First World War. He was in the trenches when he heard that he and his father had been awarded the 1915 Nobel Prize for Physics; that news reached him shortly after that of the death of his brother Robert. At only 25, he was (and still remains) the youngest ever recipient of a Nobel Prize.

Technical advances between the wars enabled scientists working in this new discipline to solve the structures of rather more complex molecules. Kathleen Lansdale, one many women who began their research careers as the Braggs' students, solved the structures of benzene derivatives and was the first to see that aromatic rings were flat. And two later developments paved the way for the explosion in structural science that characterised the later twentieth century. In 1934, John Desmond (J.D.) Bernal, who later became the first head of the School of Crystallography at Birkbeck (the predecessor department of our Biological Sciences) and his student Dorothy Crowfoot (later Hodgkin) obtained the first X-ray diffraction patterns from protein crystals. And in the following year Lindo Patterson developed a function that greatly simplified the mathematics involved in structure determination.

Even fifty years ago however, solving crystal structures was a long and at times tedious business. A typical crystallogaphy PhD thesis of the 1960s or 1970s would contain the structures of maybe three small or medium-sized molecules. It is now possible to generate as many in a few hours, so it is possible to see clearly how structures of molecules respond to changes in conditios such as temperature and pressure.

All these discoveries have been made possible by advances in technology, and particularly by the development of synchrotron radiation as a source of powerful beams of X-rays. Synchrotron radiation is produced when charged particles are accelerated radially, and synchrotrons built primarily as X-ray sources were first built in the 1980s. The UK's synchrotron, Diamond at Harwell in Oxfordshire, is currently the fifth largest in the world. It has 23 separate "beamlines", each providing a beam of X-rays with properties that have been optimised for a particular experimental technique.

Synchrotrons provide facilities for solving structures from single crystals of large and small molecules, including, of course, proteins, and from micro-crystalline samples (the latter technique is known as powder diffraction). Although structural biology attracts much of the attention (see almost all the other posts on this blog) structures of smaller molecules can still provide important insights. Sandy Blake, a crystallographer at the University of Nottingham, is using Diamond beamlines to solve the structures of novel materials called metal-organic frameworks or MOFs that are able to store gases including hydrogen (which is a potential fuel source) and greenhouse gases.

At 100, crystallography is still a young discipline but it has radically transformed many other areas of science and, through them, the world we inhabit today. This has been reflected in decisions made by the Nobel committees over the decades. The International Union of Crystallography maintains a list of Nobels awarded for ‘achievements directly related to, or involving the use of, crystallography’. There are now 29 of these, and the latest year with no crystallography-related Nobel was 2008. Even the 2013 Chemistry prize, awarded to Martin Karplus, Michael Levitt and Arieh Warshel, appears on the list: their discipline of computational chemistry would be impossible without structural knowledge obtained through crystallography.

And it almost goes without saying that protein structure, and structural biology more generally - the disciplines taught in this course and its associated MSc - owe their existence to the development of X-ray crystallography.

This blog post is based on an article I wrote for the Royal Society of Chemistry's membership journal, Chemistry World. It will be published in the January 2014 issue of the journal.

Welcome to PPS students 2012-13!

This post is very like one I wrote at exactly this time last year. This is because what I have to say now is very, very similar...

I would like to offer a warm welcome to the Principles of Protein Structure blog to all students who have just started studying Birkbeck's Principles of Protein Structure course!

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. Throughout the taught course (but more often in the later part of the course) I will post reports of recent developments. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The overall title of the programme for Autumn 2012 is "Molecular Mechanisms of Neurodegeneration": a fascinating topic which relates quite closely to some of the material we cover in the later sections of the course. Other posts may be reports from conferences or summaries of recently published papers in protein structure, protein bioinformatics and allied areas.

Some of the posts on this blog are written by "guest blogger" Jill Faircloth, who took the MSc in Structural Molecular Biology a few years ago and is now working as a freelance science communicator. She introduces herself in this post written in March 2012, in which she also describes how she found the later part of the PPS course and her thoughts on the two choices available for the second year of the MSc.

Do, if you get a chance, look through some of the earlier blog posts to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

And the best of luck for the 2012-13 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom
Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure

Welcome to new students!

A warm welcome to the Principles of Protein Structure blog to all students who have just started studying Birkbeck's Principles of Protein Structure course!

I run this blog to link the material that you will be studying in the course to new research developments in the areas of protein structure and function and related aspects of biotechnology and medicine. Throughout the taught course (but more often in the later part of the course) I will post reports of recent developments. I might, example, report on talks given in the ISMB seminar series run jointly by the Department of Biological Sciences at Birkbeck and research departments in neighbouring University College London. The overall title of the programme for Autumn 2011 is "“Proteins of the Future: Mechanism, Evolution and Design” which is closely connected to the content of the PPS course. Other posts may be reports from conferences or summaries of recently published papers in protein structure, protein bioinformatics and allied areas.

Do, if you get a chance, look through some of the earlier posts on the blog to see the kind of topics that we will be discussing. However, don't be discouraged if at this stage of the course you find the science presented there difficult to understand. I can assure you that it will get easier!

And the best of luck for the 2011-12 PPS course and for your studies at Birkbeck! We hope that many of you will go on to complete our MSc in Structural Molecular Biology.

Best wishes,

Dr Clare Sansom

Senior Associate Lecturer, Biological Sciences, Birkbeck and Tutor, Principles of Protein Structure