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Online Laboratory Magazine
07/25/2021

04/27/2018

European funding for improved efficiency of single particle imaging experiments

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With Single Particle Imaging (SPI) techniques scientists use intense X-ray light sources to produce 3D visualizations of the atomic structure and dynamics of biological molecules such as viruses and proteins. As the name suggests, the goal is to only have to use a single molecule, thereby circumventing the need for sample preparation steps such as crystallization. This approach is particularly interesting for biological systems that are notoriously difficult to crystallize such as very large or flexible protein complexes and membrane proteins.

Recent advances in SPI techniques, and the improvements in technologies available, such as the intense X-ray light sources at European XFEL, mean scientists can now produce 3D models of large molecules such as viruses. Furthermore, the availability of highly sensitive detectors means it is also possible to monitor functional cycles of large molecular assemblies in time-resolved experiments. However, SPI experiments are still a compromise between either producing static structures which require a lot of sample and data, or coarse grained time-resolved data with high levels of background noise and a lack of structural detail.

Dr. Charlotte Uetrecht, junior group leader at the Heinrich Pette Institute, Leibniz Institute for Experimental Virology, in Hamburg, and European XFEL guest scientist within the sample environment group, aims to address some of these drawbacks. Together with international collaborators she plans to develop a prototype mass spectrometer to be integrated into a prototype experiment chamber at the SPB/SFX instrument at European XFEL. The new mass spectrometer will separate biomolecules in terms of mass and conformation, thereby increasing the efficiency of SPI experiments. "An important aspect of the project is the orientation of protein complexes upon imaging" said Uetrecht. "This will result in big advantages for the data analysis and acquisition" she adds. The project - "MS SPIDOC" - is funded with 3.7 million Euros within the EU's Horizon 2020 "H2020-FETOPEN-2016-2017" (Research and Innovation Action). The Heinrich Pette Institute coordinates the project (801406) which will run for three years from September 2018.

Uetrecht, whose research interest focusses on using mass spectrometry for structural biology, was also recently awarded a prestigious ERC starting grant of 2 million Euro by the European Research Council (ERC). During the five year project, Uetrecht and her team will develop a novel mass spectrometry system based on recent developments in the fields of native mass spectrometry, ion mobility and fragmentation techniques. In particular, the project, entitled "Spock's MS" (759661), will explore the potential of soft X-ray free-electron lasers for fragmentation experiments. A software pipeline for analysis and data processing will also be developed.

Uetrecht's research at European XFEL is based on previous work done at the free-electron laser FLASH at DESY. "At FLASH we developed a sample delivery system using mass spectrometry for single particle imaging. By hitting protein ions with the X-rays inside the mass spectrometer we were able to detect the resulting fragments in the inbuilt time of flight," explains Uetrecht. "The results were promising and so we planned to investigate the ability of free-electron lasers as high intensity light sources for fragmentation of protein complexes all the way down to small particles such as peptides."

Source: European XFEL GmbH