New protein crystal harvester on MASSIF-1
A new, EMBL-developed CrystalDirect® harvester has been integrated into the macromolecular crystallography beamline MASSIF-1 (ID30A-1), offering users a unique and fully automated ‘plate-to-beam’ service for crystals – from crystal growth to data collection and structure determination.
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Macromolecular crystallography is one of the techniques used to determine the 3D structures of macromolecules, such as proteins, by directing X-rays at regularly structured arrays of molecules packed together in a crystal. The whole process involves several manual steps that include growing, harvesting and cryo-cooling the crystal, before using it to perform X-ray diffraction experiments at a synchrotron beamline. Thanks to technological innovations, some of these steps, such as data collection, data processing and structure determination, have been progressively automated over the years.
Already considered one-of-a-kind for being the only fully automated synchrotron beamline dedicated to structural biology in the world, MASSIF-1 (also known as ID30A-1), jointly operated by the ESRF and the EMBL Grenoble, delivers high-throughput, reliable data collection and crystal characterisation capabilities for academic researchers and the pharmaceutical industry. However, until recently, harvesting crystals remained a manual undertaking, using a microscope to ‘fish out’ crystals often only a few microns in size, and could still constitute an obstacle for certain projects as the harvesting and cryo-cooling of samples of protein crystals could turn out to be impossible or detrimental to the diffraction quality.
To automate the crystal-mounting bottleneck, the EMBL Grenoble Instrumentation and High Throughput Crystallography (HTX) teams developed a robot that can automatically mount crystals by cutting them out of the drops they are grown in using a laser. The robot, called the CrystalDirect® harvester, has been used at the HTX platform for many years to automatically harvest and cryo-cool crystals for the platform’s users. In order to further streamline and automate this process, a collaboration of scientists and engineers from the ESRF Structural Biology group and the EMBL Grenoble have now integrated a new CrystalDirect® harvester in the experimental hutch of the beamline (Figure 1).
Fig. 1: The CrystalDirect® high precision robot installed in the MASSIF-1 experimental hutch. Image: Stuart Ingham/EMBL
The EMBL CrystalDirect® technology automatically harvests the crystal from the crystallisation plate where it has been grown, and preserves the crystal sample by cryo-cooling it. A robotic arm then collects the crystal samples from the CrystalDirect® harvester and directly positions them on the microdiffractometer goniometer head, ready for data collection. This enables a complete automation of the crystal delivery process, from crystals grown on plates to data collection and structural determination at the beamline (i.e., ‘from plate to beam’).
During the process of integrating the CrystalDirect® harvester into the beamline, the teams worked on fine-tuning the processes to speed them up and increase the overall throughput. Several months of testing the new configuration, pipelines and processes took place during the day, while regular data collection was scheduled at night thanks to the beamline’s automated workflows. The setup of this new configuration is now complete, with the beamline able to harvest crystals and collect data simultaneously. New experimental modalities are also available, such as high-throughput, room-temperature data collection for a better understanding of protein dynamics, and automated crystal dehydration experiments to improve crystal quality parameters.
This innovative, state-of-the-art instrument for structural biology, now open to users, will further streamline operations and open new experimental opportunities.
Further information can be found at: https://www.embl.org/news/lab-matters/two-in-one-combining-massif-1-and-crystaldirect/
About the beamline: MASSIF-1 |
MASSIF-1 is a unique facility for the high-throughput, fully automatic characterisation and data collection of crystals of macromolecules. The service is not designed to replace user visits to the synchrotron but rather to do the hard work of screening crystals or collecting data sets through the night, freeing researchers to spend time on more challenging data collection problems and study the underlying biology. Beam time is booked flexibly and samples then enter a queuing system. Users interact with the beamline by describing experimental requirements, which are used by the beamline software to set data collection parameters in a database, ISPyB, where results are also viewed and downloaded. The service is made possible using the latest automation, a highly intense X-ray beam (5 x 1012 ph/sec in a flexible spot between 100 and 10 µm diameter) and complex workflows that fully evaluate samples, centre the best volumes and collect diffraction data sets optimised for maximum resolution with minimised radiation damage. |