The last few years have seen dramatic changes and developments in Macromolecular Crystallography (MX) synchrotron beamlines to counter the demand from the user community. High throughput techniques for protein expression, purification and nano-scale crystallisation have emerged and concurrently with this, more and smaller crystals are increasingly brought to the synchrotron. As a consequence, the reliability and performance of beamlines as well as the sample environment have been optimised towards an ergonomic environment and miniaturisation with high levels of automation.
Data collection at the ESRF
In this context, the ESRF, the EMBL and the UK-MRC, in collaboration with other European centres, are developing tools to make MX data collection fully automated.
• Sample freezing and shipping
A European standard for sample holders and shipping of samples to the synchrotron has been defined within the SPINE1 project. Users are encouraged to use the SPINE standard sample holders and the SPINE sample pucks to take full advantage of the beamline automation devices.
• Information flow from home lab to the beamlines
The information and identification related to every sample is provided to the beamline information system software ISPyB2 that is accessible from the user home laboratory via a web browser. Users are able to feed in information useful to their beamline experimental time.
• Data collection
The users, assisted by a software expert system (DNA3), perform sample screening and evaluation . From the pre-processed results, users choose an appropriate data collection strategy.
Programme of the course
A 5-day week of training centred on these latest beamline technological developments will be organised in Grenoble from the 9 to the 13 October 2006. A mixture of lectures, tutorials and practical sessions will cover bio-informatics, robotic protein production and crystallisation, as well as structure solution using data collected on synchrotron MX beamlines.
Day 1: the week will start with one full day of courses and tutorials on the latest bio-informatics tools for protein sequence analysis. Students will learn how to extract a maximum of structural information and data to aid expression and crystallisation from primary sequences. During Day 2 high-throughput techniques for protein expression, purification and crystallisation will be presented and a guided tour of the local robotic facilities will be organised. Day 3: after an overview course of the principle biophysical techniques to characterise biological macromolecules, protein crystallography methods will be introduced in very practical terms. Students will start a two-days training on the ESRF protein crystallography beamline environment described above (Days 3 to 5). The training week will end by one day (Day 5) of phasing tutorials (using a tutor test data or the student’s own data collected on the beamlines), model building and refining.
Students are expected to bring either:
- Their own protein sample in solution and will have the opportunity to screen crystallisation conditions using a nano-litre robot or
- Their own cryo-cooled crystals mounted on SPINE standard sample holders to screen them on the ESRF beamlines.
They will be trained to use the ISPyB system for sample identification and shipment, as well as in the use of the beamline sample changer and automated data collection system (DNA).
Protein robotic production and purification
Protein robotic crystallisation and observation
Data collection on heavily automated beamlines
Data reduction and phasing
David Hall and Edward Mitchell, ESRF
Martin Walsh, MRC
Hassan Belrhali, EMBL
Nadine Petricola, ESRF (HSC2 secretary)