Expanding structural proteomics to monitor proline isomerization

SUMMARY The correct cis/trans amide bond configuration at prolines is vital for protein folding and altered configurations are associated with the onset of several diseases. At present, determining the cis/trans state of prolines necessitates either high-resolution structural determination or extensive NMR experiments, which are often not feasible for larger or more complex protein assemblies. For this reason, it is highly desirable to develop new analytical tools to enable direct identification of proline cis/trans states across a protein sequence. We propose development of tandem proteolysis applying a combination of cold liquid chromatography, proline specific proteases, ion mobility mass spectrometry, and ultraviolet photodissociation (UVPD) to resolve and directly monitor proline configurations across a protein sequence. Integrated with existing bottom-up proteomics pipelines, this approach will provide a much-needed tool for analyzing proline cis/trans isomerization that is capable of handling sample complex samples and far faster than current methods. Project Number: 1R01GM164435-01 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Miklos Guttman | Institution: UNIVERSITY OF WASHINGTON, SEATTLE, WA | Award Amount: $1,368,400 | Activity Code: R01 | Study Section: Enabling Bioanalytical and Imaging Technologies Study Section[EBIT] View on NIH RePORTER: https://reporter.nih.gov/project-details/11339873