Development of the method of X-ray footprinting at the Advanced Light Source

May 20, 2020

Development of the method of X-ray footprinting at the Advanced Light Source

The method of X-ray footprinting mass spectrometry (XFMS) is a relatively recent structural biology characterization method that was pioneered at the National Synchrotron Light Source at Brookhaven National Laboratory in the late 90's. XFMS is an in situ hydroxyl radical (•OH) labeling method; X-ray irradiation dissociates solvent water to produce hydroxyl radicals, which covalently modify side chains of proteins which are solvent accessible.

More specifically, residues which are in proximity to water molecules (either bulk or bound) are modified to a greater extent than residues which are not in proximity to water. Because liquid chromatography-mass spectrometry is then used to analyze the stable covalent modifications produced, the data provide a “water map” at the single residue level, which is then used to determine sample conformation.

Since its introduction, the use of XFMS to investigate structural features and conformational changes of macromolecules in the solution state has grown substantially and has been successfully applied to systems ranging from single domain proteins to in vivo ribonucleoprotein assemblies. The method is highly complementary to the more widely used structural elucidation techniques for biological macromolecules such as crystallography, HDX, and cryo-electron microscopy. This talk will describe the XFMS method, its advantages and disadvantages relative to other methods, its implementation at the ALS, and some recent instrumentation upgrades to make the method more accessible to a wider user population.

Speaker: Corie Ralston holds a B.S. in Physics from the University of California at Berkeley, and a Ph.D. in Biophysics from the University of California at Davis. She completed a post-doctoral fellowship at Brookhaven National Laboratory during which she helped develop the method of X-ray footprinting as a structural investigation technique for proteins and nucleic acids. She is currently a Biophysicist Staff Scientist in the Molecular Biophysics and Integrated Bioimaging division at Berkeley Lab and the Interim Director of the Biological Nanostructures facility at the Molecular Foundry. At the Advanced Light Source, she is developing high-throughput methods for X-ray footprinting.