Solving Protein Structures from Scratch

A novel tool to determine the structure of difficult to crystallize proteins.

Image courtesy of Max Planck Society
The weak signal of two gadolinium ions - shown as brilliant spheres - was used to solve the high resolution crystal structure of the model protein lysozyme.

The Science

Using the Linac Coherent Light Source x-ray free-electron laser, an international team of researchers generated a complete 3-dimensional model of the protein lysozyme without any prior knowledge of its structure. The emerging technique of serial femtosecond crystallography provides a potential new avenue to study important biomolecules with unknown structure.

The Impact

This proof-of-principle demonstration opens a path to determining, from scratch, biological structures that form crystals too small for analysis with conventional X-ray sources. Adapting and refining the technique will allow more complex proteins to be examined such as membrane proteins and their complexes and potentially provide new targets for drug development.


The determination of protein crystal structures is hampered by the need for large, macroscopic crystals. X-ray Free-Electron Lasers (FELs) provide extremely intense X-ray pulses of femtosecond duration, which allow data collection from nano-to-micrometer-sized crystals without the limitations set by radiation damage common for traditional approaches. FELs can produce high resolution, undamaged structural information; however, all of these protein structure determinations were based on related, previously known structures. Using the Linac Coherent Light Source, an international team of researchers showed X-ray FEL data can be used for de novo protein structure determination, i.e., without prior knowledge about the structure. Micron-sized protein crystals were introduced in the FEL beam using a liquid microjet, exposing one crystal after the other to collect snapshot diffraction data in a serial fashion. Using this emerging technique of serial femtosecond crystallography, single-wavelength anomalous scattering measurements were performed on the well-established model system lysozyme, in complex with a lanthanide compound. Experimental phases were determined by stitching together the information of 60,000 diffraction snapshots, resulting in an experimental electron density map good enough for automated building of the protein structure. This result shows the feasibility of determining novel protein structures using FELs. It is anticipated that serial femtosecond crystallography will become an important tool for the structure determination of proteins that are difficult to crystallize such as membrane proteins which are important drug targets.


Ilme Schlichting
Max Planck Institute for Medical Research, Heidelberg, Germany


Portions of this research were carried out at the Linac Coherent Light Source, a DOE Office of Science  User Facility operated by Stanford University and supported by DOE Office of Science Basic Energy Sciences program. The CXI instrument was funded by the LCLS Ultrafast Science Instruments (LUSI) project funded by the DOE Office of Science Basic Energy Sciences program. We acknowledge support from the Max Planck Society and from the EU for an Incoming Scientist Award to R.B.D.


Thomas R.M. Barends, Lutz Foucar, Sabine Botha, R. Bruce Doak, Robert L. Shoeman, Karol Nass, Jason E. Koglin, Garth J. Williams, Sebastien Boutet, Marc Messerschmidt, Ilme Schlichting. “De novo protein crystal structure determination from X-ray free-electron laser data.” Nature 505, 224 (2014). [DOI: 10.1038/nature12773]

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