A mixing flow-cell for difference SAXS studies of protein conformational changes.

We aim to observe structural perturbations using small angle x-ray scattering (SAXS) for two different ion channels (KcsA and LGC-39) and in cytochrome P450. We will take advantage of 3D printing technologies to overcome previous limitations in difference X-ray scattering studies when mixing samples. A recent proof-of-principle test at ID02 of the ESRF successfully observed structural changes in KcsA by using UV light to release caged protons, thereby jumping the sample from pH 7 (neutral) to 4 (acidic). We will extend this study using a mixing cell to induce a similar pH jump. Should both approaches yield similar structural changes, the same approach will probe ligand-binding-induced conformational changes in a ligand gated ion channel (LGC-39) isolated from the model-organism C. elegans, and ligand-binding-induced conformational changes in cytochrome P450, a drug metabolizing enzyme. This work increases our understanding of neuronal transmission selectivity and enzyme activity.