The Beamline X28C of the Center for Synchrotron Biosciences: a National Resource for Biomolecular Structure and Dynamics Experiments Using Synchrotron Footprinting

Structural mapping of proteins and nucleic acids with high resolution in solution is of critical importance for understanding their biological function. A wide range of footprinting technologies have been developed over the last ten years to address this need. Beamline X28C, a white‐beam X‐ray source at the National Synchrotron Light Source of Brookhaven National Laboratory, functions as a platform for synchrotron footprinting research and further technology development in this growing field. An expanding set of user groups utilize this national resource funded by the National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health. The facility is operated by the Center for Synchrotron Biosciences and the Center for Proteomics of Case Western Reserve University. The facility includes instrumentation suitable for conducting both steady‐state and millisecond time‐resolved footprinting experiments based on the production of hydroxyl radicals by X‐rays. Footprinting studies of nucleic acids are routinely conducted with X‐ray exposures of tens of milliseconds, which include studies of nucleic acid folding and their interactions with proteins. This technology can also be used to study protein structure and dynamics in solution as well as protein–protein interactions in large macromolecular complexes. This article provides an overview of the X28C beamline technology and defines protocols for its adoption at other synchrotron facilities. Lastly, several examples of published results provide illustrations of the kinds of experiments likely to be successful using these approaches.