Project summary: The MacCHESS Synchrotron Source for Structural Biology facilitates the utilization of both established and emerging technologies to enable the structural characterization of proteins involved in critically important biological processes with significant consequences for diseases such as cancer, bacterial and viral infections, and neurodegenerative disorders. Recent upgrades to CHESS, including improvements to the storage ring and newly designed beamlines have provided state-of-the-art facilities and enabled greater capabilities since June 2019. MacCHESS will continue to support more than 100 investigator projects, funded by NIH and other government institutions, through two major Technology Operations Cores. These are: 1) Facility for Flexible Crystallography, which will take advantage of unique MacCHESS capabilities to enable the development of new X-ray techniques that may be used to broaden our knowledge of essential biological processes. Examples include continued development of methods for room temperature crystallography toward identifying new structural or ligand/drug binding states, the application of high pressure to crystals, and analysis of macromolecular motions through the study of X-ray diffuse scattering. A high level of support for more routine macromolecular crystallography will also be provided. These methods will help to provide important new insights including the binding interactions and conformational transitions that cell signaling proteins and drug targets undergo which are necessary for their function, and the macromolecular motions essential for the catalytic activities of enzymes that regulate key metabolic processes, DNA transcription and repair, and different aspects of RNA biology. 2) Facility for Biological Small Angle X-ray Scattering (BioSAXS), which will implement state-of-the-art hardware, software, and expertise to support the BioSAXS technique that continues to be in high demand. In addition to determining the shapes of proteins, nucleic acids, and larger protein assemblies in solution, BioSAXS allows researchers to obtain information regarding global conformational changes within macromolecular complexes (e.g., membrane receptors, RNA-splicing complexes, large or multi-subunit enzymes) and the changes in their oligomeric states that have important functional consequences. This core will also provide the necessary equipment and expertise for investigators interested in performing time resolved BioSAXS or BioSAXS studies conducted under high pressure. MacCHESS will provide a strong Administration Core to support these activities and will continue to educate and help to train users and investigators from the biomedical research community new to the field of structural biology, through a Training and Outreach Core. Collectively, these efforts will offer unique opportunities to our users for pursuing some of the most challenging questions in structural biology and for obtaining structure-function...