Optimisation of crystal growth using a microfluidic technology-based crystallisation bench (ESR1)

Applications are invited for an Early Stage Researcher (ESR)/PhD studentship positions funded by The Marie Skłodowska-Curie Actions (MSCA) Innovative Training Networks (ITN) programme “Rationalising Membrane Protein Crystallisation” (RAMP) and based in the Synchrotron Group at the Institut de Biologie Structurale (IBS), France. The Institut de Biologie Structurale (IBS) is a research centre for integrated structural biology funded by the CEA, the CNRS and the Université Grenoble Alpes (UGA).

Over the last 20 years, and while the crystallisation of membrane proteins remained almost entirely empirical, a combination of experimental and computational work greatly advanced the understanding of the nucleation and growth of crystals in many other systems, from ice to globular proteins. Here we aim to build on this work to apply predictive models and microfluidic techniques to membrane protein crystallisation to tailor crystal sizes and morphologies required for different downstream structure determination approaches. This PhD project is one of 12 in the RAMP training network. Other PhD students in the RAMP network will develop advanced methods for crystallisation and modelling of the membrane protein crystallisation process and apply it to the structure determination of important membrane proteins.

This project focuses on specific challenges for the crystallisation of membrane proteins in which the precise control of the crystal size and morphology is essential. Because neutrons interact very weakly with matter, neutron crystallography require large crystals, with volumes of > 0.01 mm3. This is the requirements of neutron sources such the ESS (European Spallation Source in Lund), the site of the new generation neutron source in Europe. At the other extreme are the requirements of serial crystallographic methods that are being increasingly used at synchrotron sources (serial synchrotron crystallography) due to advances in micro- and nano- focus beamlines, as well as at rapidly developing ultra-bright free-electron laser sources (serial femtosecond crystallography) at the Hamburg Centre for Ultrafast Imgaging. These new X-ray sources make possible the structure determination of previously intractable proteins, where uniform population of small microcrystals in the 1-20 μm size is required.

Objective of the PhD project is to further develop and adapt the current crystallization bench (Junius et al., 2016, J. Applied Cryst., 49, 806-813) to the crystallization of membrane proteins for both, neutron and time-resolved X-ray crystallography. The existing flow-cell dialysis set-up combines the control of the temperature and of the chemical composition of the crystallization solution. The technological goal of the PhD student will be to re-design the temperature-controlled fluidic device to successfully integrate in situ UV spectroscopy. An important part of the work will be devoted to exploring phase behavior in lipid-based multicomponent systems and using statistical modeling to rationally improve the final stages of crystallization of membrane proteins, especially when crystallization conditions have been obtained but where the crystals are too small or badly ordered. The new instrument will be used to grow crystals of model membrane proteins with sizes and morphologies required for different downstream structure determination approaches and the crystal perfection of obtained crystals will be evaluated. The project will involve secondments to other network partners with specialised membrane protein crystallisation and computational/theoretical modelling expertise such as University of Leeds, University of Aarhus, University of Surrey and University of Maynooth, and cosupervision of NatX-ray.

This project will involve crystallisation and X-ray and Neutron diffraction analysis of crystals of model membrane proteins, as well as instrumentation, automation and methodological development in protein crystallography, with support from the existing expertise in the group. Crystals will be studied at national and international synchrotrons, neutron sources and free electron lasers.

Applicants must have the background and expertise required for the position, be in the first four years full-time equivalent research experience of research career, since, e.g., completion of their masters’ degree, do not already possess a doctorate degree, be willing to move to FRANCE and to other countries within the network, they must also be eligible for a working permit for FRANCE in which they have not lived for more than 12 months over the last 3 years (short stays, such as holidays, are not taken into account), be proficient in both written and spoken English.

Candidates must comply with EU and Université Grenoble Alpes eligibility criteria. Due to the EU rules to promote mobility, you are not eligible for a position in a country where you have lived (worked, studied) for more than 12 months in the last 3 years. So for this position you are eligible, unless you have studied or worked in France for more than 12 of the last 36 months. For applicants finishing or who have just finished their degree, this typically means that you can be graduating from any university except a French university.

Further details on this project are available from Dr. Monika Spano (monika.spano@ibs.fr).

Author: Richard Sear

Computational physicist at the University of Surrey. My research interests are in COVID-19 transmission, especially masks, soft matter & biological physics

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