07/30/2022: 5:30 PM - 7:30 PM
Portland Marriott Downtown Waterfront
Room: Exhibit Hall
Phosphoenolpyruvate carboxykinase (PEPCK), an essential enzyme that converts oxaloacetate to phosphoenolpyruvate and gates the gluconeogenesis pathway has recently been found to be upregulated in certain cancers.(1,2) Utilizing MMQX (Millisecond Mix-and-Quench Crystallography) we collected time-resolved crystallography data at timepoints of 40ms, 120ms, and 200ms. These datasets were able to capture PEPCK motions associated with substrate binding and catalysis as well as binding positions of the phosphoenolpyruvate and carbon dioxide products.(3) In addition to time-resolved crystallography, we also performed multi-temperature crystallography of PEPCK to better understand the energy landscape in steady-state conditions. These experiments captured the opening of the omega active site gating loop in PEPCK. Taken together, these experiments greatly improve our understanding of PEPCK's structural fluctuations.
1. Johnson, T. A. & Holyoak, T. The ω-loop lid domain of phosphoenolpyruvate carboxykinase is essential for catalytic function. Biochemistry 51, 9547–9559 (2012).
2. Leithner, K., Hrzenjak, A., Trötzmüller, M., Moustafa, T., Köfeler, H. C., Wohlkoenig, C., Stacher, E., Lindenmann, J., Harris, A. L., Olschewski, A. & Olschewski, H. PCK2 activation mediates an adaptive response to glucose depletion in lung cancer. Oncogene 2015 348 34, 1044–1050 (2014).
3. Clinger, J. A., Moreau, D. W., McLeod, M. J., Holyoak, T. & Thorne, R. E. Millisecond mix-and-quench crystallography (MMQX) enables time-resolved studies of PEPCK with remote data collection. IUCrJ 8, 784–792 (2021).