A genetically encoded anomalous SAXS ruler to probe the dimensions of intrinsically disordered proteins

• DOI: 10.1073/pnas.2415220121
• Publication Date: 2024-12-06T18:42:57Z
• Authors: Miao Yu; Andrey Yu. Gruzinov; Hao Ruan; Tom Scheidt; Aritra Chowdhury; Sabrina Giofrè; Ahmed S. A. Mohammed; Joana Caria; Paul F. Sauter; Dmitri I. Svergun; Edward A. Lemke
• Keywords: Small-angle X-ray scattering; Intrinsically Disordered Proteins; Radius of gyration; Small-Angle Scattering

Intrinsically disordered proteins (IDPs) adopt ensembles of rapidly fluctuating heterogeneous conformations, influencing their binding capabilities and supramolecular transitions. The primary conformational descriptors for understanding IDP ensembles—the radius gyration ( R G ), measured by small-angle X-ray scattering (SAXS), the root mean square (rms) end-to-end distance E probed fluorescent resonance energy transfer (FRET)—are often reported to produce inconsistent results regarding expansion as a function denaturant concentration in buffer. This ongoing debate surrounding FRET-SAXS discrepancy raises questions about overall reliability either method quantitatively studying properties. To address this discrepancy, we introduce genetically encoded anomalous SAXS (ASAXS) ruler, enabling simultaneous direct measurements without assuming specific structural model. ruler utilizes noncanonical amino acid with two bromine atoms, providing an signal precise measurements. Through approach, experimentally demonstrate that ratio between varies under different denaturing conditions, highlighting intrinsic properties IDPs source observed SAXS-FRET rather than shortcomings established methods. developed ASAXS emerges versatile tool both folded proteins, unified approach obtaining complementary site-specific information experiments, thereby contributing deeper protein functions.