Page 1 ISMB NEWS – Issue 10 – Summer 2014

ISMB NEWS

Issue 10, Summer 2014

ISMB NEWS is a news bulletin published in order to share information between the various departments involved in the Institute of Structural and Molecular Biology. This includes information such as events, research highlights, new staff appointments, awards and grants.

Your comments, suggestions and contributions are welcome and will help us put together a newsletter which meets your expectations. Please email the ISMB administrator at ismb-admin@ismb.lon.ac.uk.

In this issue WELCOME TO… FAREWELL TO…

IN FOCUS: ISMB PROFILES

Dr Kristine Arnvig Dr Filipe Cabreiro RESEARCH HIGHLIGHTS

Dr Carolyn Moores ISMB Featured articles and commentaries STUDENT NEWS

UPDATE ON CENTRES & LABORATORIES

AWARDS, PRIZES & GRANTS UPCOMING EVENTS

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WELCOME TO… FAREWELL TO…

We are pleased to welcome Prof Geraint Rees, Director of the UCL Institute of Cognitive Neuroscience, who has been appointed to the next Dean of the Faculty of Life Sciences from 1st September 2014. Farewell to Prof Mary Collins, the departing Dean who has led the faculty for the last 5 years.

We welcome the following Post-Doctoral Research Associates who have recently joined ISMB lab groups: Drs Vincent Gray and Matthias Glaser (Dr Erik Arstad’s group), Dr Susan Andrew (Dr Richard Hayward’s group), Dr Tanya Singh (Prof David Jones’ group), Drs Paul Ashford, Jacek Grzbyta and Aurelio Moya Garcia (Prof Christine Orengo group), Dr Katja Ota (Prof Helen Saibil’s group), and Dr Juliana Sakamoto Yoneda (Prof Bonnie Wallace’s group).

Farewell to the following Post-Docs: Drs Alison Cuff, Tim Nugent and Sarah Tarr.

We bid farewell to Prof Christine Slingsby, retiring Professor of Structural Biology and Deputy Head of Research in the Department of Biological Sciences, Birkbeck, and ISMB Executive Committee member. Christine made a remarkable contribution to Birkbeck’s department of Biological Sciences (formerly School of Crystallography) over many years – she will be missed and is wished the very best in her retirement.

Structure of a Type IV Secretion System

(Low et al (2014) Nature 508:550-553)

Page 2 ISMB NEWS – Issue 10 – Summer 2014

IN FOCUS: ISMB PROFILES

Dr Kristine Arnvig, Lecturer, Research Department of Structural and Molecular Biology, UCL

Dr Kristine Arnvig is a Lecturer in molecular microbiology in the ISMB. Kristine works with regulatory RNA in Mycobacterium tuberculosis, the bacterium that causes tuberculosis in humans. This disease is still a major threat to global health claiming around 1.5 million lives every year with a steadily increasing number of deaths being caused by multidrug resistant strains of M. tuberculosis. The bacterium thrives and persists in the human host as multiple subpopulations, which reflect the ability for significant changes in gene expression. Kristine has been working with regulatory RNA in M. tuberculosis for many years; initially with antitermination of

ribosomal RNA transcription, and currently with small RNAs and riboswitches. Small RNAs are small trans-acting molecules that can regulate protein activity by sequestration, or mRNA stability and/or expression by basepairing. They are often associated with stress and changes in the environment and this is also the case for many M. tuberculosis small RNAs. The RNA research focuses on how and when these regulators are expressed and degraded and what other molecules they interact with in the cell, and ultimately, what role they play in the survival and adaptation of M. tuberculosis to the host environment.

Riboswitches are RNA elements that are part of the mRNAs they regulate. These elements regulate expression of their cognate mRNA upon binding a small molecule ligand. For both types of ribo-regulation the overall aim is to get a further insight into post-transcriptional regulation of gene expression in M. tuberculosis and to integrate this with the well-established protein-based regulatory network in M. tuberculosis.

[Details of work and papers at https://iris.ucl.ac.uk/iris/browse/profile?upi=KARNV52] [email: k.arnvig@ucl.ac.uk]

Dr Kristine Arnvig

Dr Filipe Cabreiro, Sir Henry Dale Fellow and Lecturer, Research Department of Structural and Molecular Biology, UCL and Birkbeck

Filipe is a lecturer in metabolism and metabolic disease in the ISMB. He obtained his PhD degree in Biochemistry at the Université Paris Diderot in France, where he exploited the biological mechanisms underlying molecular stress protection and zinc homeostasis (Cabreiro et al., JBC 2008; Cabreiro et al., Antiox redox signal. 2009). In 2008, Filipe joined the group of Prof David Gems within the Institute of Healthy Ageing (IHA)/GEE at UCL as a post-doctoral researcher. Since then, he has broadened his background knowledge in several key areas such as genetics, molecular biology, metabolism and microbiology. He undertook several projects simultaneously to investigate different aspects of ageing. These topics included the role of antioxidant

metabolism and signaling in ageing (Cabreiro et al., Free Radic. Biol. Med. 2011), the role of sirtuins in ageing and disease (Cabreiro et al., Nature 2011), and the role of the anti-diabetic drug metformin in C. elegans metabolism and ageing (Cabreiro et al., Cell 2013). More recently he has pioneered the use of the model organism C. elegans to study how drug-microbiota interactions affect host metabolism and ageing. This work has led to developing a focus on host-microbiota interactions and metabolic disorders (Cabreiro and Gems, EMBO Molec. Med. 2013). He is currently a Sir Henry Dale Fellow at the ISMB having been awarded a prestigious fellowship from the Royal Society/Wellcome Trust to explore the gut microbial action of metformin. Using a combination of metagenomics and gnotobiotics, his new research aims to gain insight into the gut microbial action of drugs in higher organisms and to develop strategies for targeting the microbiota to treat host metabolic disease.

Filipe has an active interest in public engagement. He was an active member in the organisation of the successful “Healthy Ageing” stand organised by the IHA, UCL for the Summer Science 2012 festival at the Royal Society, and at Bangkok Science and Technology Fair 2012, which is open to schools and the general public to inform people on how diet affects ageing and how can we study these effects using simple and tractable model organisms. More recently, Filipe has participated and interviewed for a podcast for Radio Cardiff's science show, Pythagoras Trousers on the role of the microbiome in health.

[Details of work and papers at https://iris.ucl.ac.uk/iris/browse/profile?upi=FGOME58] [email: f.cabreiro@ucl.ac.uk]

Dr Filipe Cabreiro

Page 3 ISMB NEWS – Issue 10 – Summer 2014

RESEARCH HIGHLIGHTS

Under the Hood of an Essential Mitotic Motor

The success of mitosis depends on the activity and integrity of the bipolar spindle, a macromolecular

machine that enables organisation and separation of duplicated DNA. The spindle is built from

microtubules (MTs), which are dynamic, polar polymers of αβ-tubulin dimers. MT dynamics are

essential for chromosome segregation, while MTs also act as tracks for molecular motors that organise

the spindle. Kinesins are a superfamily of ATP-dependent, MT-based nano-motors that play diverse

roles in cell division. They are defined by their conserved motor domains, which contain both ATP- and

MT-binding sites. Subtle modifications within these motor domains have a dramatic impact on the

properties of individual kinesins and, thus, on their cellular functions. However the relationship between

molecular variation and motor function is largely unknown.

The goal of our study was to elucidate the molecular mechanisms of the kinesin-5 mitotic motor.

Kinesin-5 motors are essential for mitosis in many organisms. They function as tetramers, with pairs of

motor domains located at either end of a dumb-bell-shaped molecule. The kinesin-5 quaternary

structure is thus perfectly designed for the motors’ proposed MT cross-linking/sliding activity in the

spindle. These motors have been of particular interest because small molecule inhibitors of human

kinesin-5 have been identified. These drugs allosterically block motor activity and do not compete with

ATP binding. By specifically inhibiting kinesin-5s, they block mitosis and are forming the basis of drug

discovery programmes for novel cancer treatments. However, despite concentrated research efforts,

our understanding of kinesin-5’s molecular mechanism is limited and key unresolved questions remain.

In particular, because the structural basis of the motor’s ATPase cycle is not known, the precise

mechanism of motor inhibition by these small molecules is still lacking, and better or alternative drugs

cannot be rationally designed.

Answering structural questions about motor mechanism is challenging because high spatial resolution is

needed to study conformations that typically last only fractions of a second. In addition, the complexes

formed by kinesins and their MT tracks are large and dynamic. We solved these challenges by

determining a sequence of structures using cryo-electron microscopy and image reconstruction. We

thereby obtained multiple snapshots of the motor and comprehensively determined the

mechanochemical cycle of the human MT-bound kinesin-5 motor domain at subnanometer resolution.

Because of the quality of our reconstructions, we were able to calculate pseudo-atomic models of the

motor at sequential steps in its ATPase cycle. Further, the nucleotide-dependent conformational

changes revealed by these structures were validated by tracking of site-specific gold-labels. Finally, with

our collaborators, we combined kinetics and motility assays with the structural studies, to conclusively

demonstrate the critical role of specific structural elements for kinesin-5 function.

(continued overleaf)

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Our work thus provides the first comprehensive structural analysis of a mitotic kinesin throughout its

MT-based ATPase cycle, revealing significant divergence from so-called “standard” models of the

kinesin mechanism. In particular, our structures yield a determination of how kinesin-dependent

variations, and particularly the kinesin-5-specific motor domain inserts, cause this divergence. This

includes the region of the motor domain where allosteric inhibitors are known to bind, shedding light on

their molecular mechanism. Blockage of mitosis through kinesin-5 inhibition could have wider

applications in disease control, for example in killing eukaryotic pathogens, and our current research is

investigating this possibility.

This work was funded by the BBSRC. Dr Adeline Goulet is now working at the Centre National de la

Recherche Scientifique and Aix-Marseille University, Marseille, France, funded by a FEBS Fellowship.

Goulet, A., Major, J., Jun, Y., Gross, S., Rosenfeld, S.S., Moores, C.A. (2014). A comprehensive structural model of

the mechanochemical cycle of a mitotic motor highlights molecular adaptations in the kinesin family. P.N.A.S. 111,

1837-1842

Dr Carolyn Moores

ISMB Featured Articles and Commentaries

A commentary on the following article is available at http://www.ismb.lon.ac.uk/news.html

H.H. Low, F. Gubellini, A. Rivera-Calzada, N. Braun, S. Connery, A. Dugeancourt, F. Lu, A. Redzej, R. Fronzes, E.V. Orlova, and G. Waksman (2014). Structure of a Type IV Secretion System. Nature. 508:550-553.

Page 5 ISMB NEWS – Issue 10 – Summer 2014

STUDENT NEWS

Laura Stoffels awarded scholarship

Congratulations to Laura Stoffels on being awarded a prestigious PhD Enterprise Scholarship from UCL Advances. The award will provide funding of £4,000 for a three-month extension to Laura's PhD to enable her to write a thesis chapter on the commercial evaluation of her research work.

Claudia Hinze awarded British Heart Foundation PhD studentship

Congratulations to Claudia Hinze, who has been awarded a 3-year PhD Studentship from the British Heart Foundation. Claudia did a 6 month internship at the ISMB in 2013 toward her MSc at the Dresden University of Technology, and will now join Dr Emmanuel Boucrot’s lab group as a PhD student.

Benjamin Lichman wins poster prize

Congratulations to Wellcome Trust PhD programme student Benjy Lichman who won the Lilley-sponsored poster award at the 3rd Multi-enzyme catalysed processes conference held in Madrid from 7- 10th April, for his poster entitled “A Two Enzyme, One substrate, One pot synthesis of (S)- Norlaudanosoline”.

Lucy Rayner paper published

Congratulations to Lucy Rayner from Prof Steve Perkins’ group, for her recent publication in the Journal of Biological Chemistry: Rayner LE, Hui GK, Gor J, Heenan RK, Dalby PA, Perkins SJ. The Fab Conformations in the Solution Structure of Human IgG4 Restricts Access to its Fc Region: Implications for Functional Activity. J. Biol. Chem. doi:10.1074/jbc.M114.572404

Page 6 ISMB NEWS – Issue 10 – Summer 2014

UPDATE ON CENTRES & LABORATORIES

Dr Chris Kay laboratory

Structural biology relies on high-resolution methodologies to reveal protein function, mis-function and disease. Advances in areas such as EPR spectroscopy in conjunction with molecular modelling are emerging as complementary approaches to X-ray crystallography, NMR spectroscopy and electron microscopy for unravelling details of cellular processes at the molecular level. EPR employs the same principles as MRI scanners, namely a combination of a magnetic field (approximately ten thousand times stronger than the earth's) and microwaves, to investigate the structures of proteins: the molecular machines that make our cells work. In order to do this, the protein is labelled with a pair of stable free radicals which behave like a magnetic compass. When two compasses are in close proximity, they feel each other's presence, and align north to south: the closer they are the larger the effect. Our experiments use microwaves to flip the pair of free radicals from being in the favourable north-south alignment to he unfavourable alignment. This allows the energy difference between the two alignments to be measured, from which we can deduce the separation between the free radicals. By attaching the free radicals at different positions we can understand how proteins change shape as they work and by labelling different proteins

we can examine how they interact with each other. For these experiments we use nitroxide spin-labels that are attached to proteins via chemical modification of native or engineered cysteine residues. As the labels have a short tether they sample a range of conformations. Hence, rather than obtaining a single distance between a pair of labels, we measure a distribution of distances. The methodology is applicable over the range 2-10 nm. Advantages of the EPR approach include: no limitation on size; no ordering requirement as experiments can be performed in (frozen) solution; identical pairs of labels are used. Examples of how EPR distance measurements have been used at the ISMB include: 1) Validation of structural models obtained by X-ray crystallography. 2) Establishing oligomerisation states in solution. 3) Assessing the effect of substrate binding on structure. 4) Determination the structure of parts of proteins invisible in X-ray structures. Researchers interested in using this approach should contact Dr Chris Kay (c.kay@ucl.ac.uk).

Bruker EPR spectrometers located in the Department of Physics & Astronomy, UCL.

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AWARDS, PRIZES & GRANTS Recently awarded grants

Funding Body Awardee/s Details

Birkbeck Department of Biological Sciences, Birkbeck

Two Birkbeck 190th Anniversary studentships, which will fund Roxana Mironska and Arundhati Maitra. They will study on the ISMB’s Wellcome Trust PhD programme from September 2014 and April 2015 respectively.

BBSRC Prof Christine Orengo

LoLa award of £474,221 for co-investigation on project ‘A Spatio-Temporal Map of the Developmental Fly Interactome’ led by Prof Simon Hubard, Manchester University. Running from 1st March 2014 to 28th February 2017.

Marie Curie Prof Christine Orengo

€309,235 awarded for project ‘Targetting Glioblastoma Signalling’ on an Intra-European Fellowship.

MRC Project Grant Dr Kristine Arnvig £472,943 awarded for project ‘The Role of Small Regulatory RNAs in Mycobacterium Tuberculosis Pathogenesis’ from 1st September 2014 to 31st August 2017.

BBSRC Dr Joanne Santini £248,379 for co-investigation on a project led by Prof Tony Cass at Imperial College. Follow-on funding awarded for 2 years from 1st May 2014.

Alexion Pharmaceuticals/ Complement UK

Prof Steve Perkins Funding a 4 Year PhD studentship.

British Heart Foundation

Dr Emmanuel Boucrot

Funding a 4 year PhD studentship.

Grants applications – upcoming deadlines

Funding body Funding opportunities Deadlines

Wellcome Trust Investigator Awards 25th July 2014

MRC Research Boards Molecular and Cellular Medicine 10th September 2014

MRC Research Board Infection and Immunity 16th September 2014

BBSRC Responsive Research Grants 4pm, 24th September 2014

EPSRC Responsive Research Grants Applications invited at any time

Wellcome Trust Small Grants Applications invited at any time

Page 8 ISMB NEWS – Issue 10 – Summer 2014

UPCOMING EVENTS

2nd UCL Symposium on the Origin of Life

The 2nd UCL Symposium on the Origin of Life will take place on Monday 21st July. Places are limited to 180. Full details, including how to register, are available on the ISMB website at http://www.ismb.lon.ac.uk/origin_of_life.html.

ISMB Seminar series, Autumn 2014/15

The Autumn 2014 seminar series, Redesigning Biological Systems - from Single Molecules to Novel Organisms will take place on Wednesdays from 1st October 2014. Seminars will take place in the Gavin de Beer lecture theatre in the Anatomy Building, UCL. Weekly details will be announced at http://www.ismb.lon.ac.uk/seminar.html.

ISMB Friday Wraps, Autumn 2014

Autumn 2014 ISMB Friday Wraps will commence on Friday 3rd October 2014, taking place in the Glaxo Wellcome seminar room B62, Birkbeck. Weekly details will be announced at http://www.ismb.lon.ac.uk/fridaywrap.html

LSBC meeting, Summer 2014

The next London Structural Biology Club (LSBC) meeting will take place on 1st July 2014, Clore B01 Lecture Theatre, Birkbeck. The LSBC can be found on Facebook for discussion and promotion of LSBC events - https://www.facebook.com/groups/396037070496925/

ISMB Retreat, 2015

The ISMB’s 6th Retreat will take place on 30th June and 1st July 2015. Further details will be announced in the new year.

News items are added each month to the ISMB news page: http://www.ismb.lon.ac.uk/news.html

Please email items of news to the ISMB Administrator (Andrew Service): ismb-admin@ismb.lon.ac.uk.