Determination of absolute intramolecular distances in proteins using anomalous X-ray scattering interferometry

Stubhan, Samuel ; Baptist, Anna V.; Körösy, Caroline ; Narducci, Alessandra; Moya Muñoz, Gustavo Gabriel; Wendler, Nicolas ; Lak, Aidin; Sztucki, Michael; Cordes, Thorben und Lipfert, Jan (2025): Determination of absolute intramolecular distances in proteins using anomalous X-ray scattering interferometry. In: Nanoscale, Bd. 17, Nr. 6: S. 3322-3330

Abstract

Biomolecular structures are typically determined using frozen or crystalline samples. Measurement of intramolecular distances in solution can provide additional insights into conformational heterogeneity and dynamics of biological macromolecules and their complexes. The established molecular ruler techniques used for this (NMR, FRET, and EPR) are, however, limited in their dynamic range and require model assumptions to determine absolute distance or distance distributions. Here, we introduce anomalous X-ray scattering interferometry (AXSI) for intramolecular distance measurements in proteins, which are labeled at two sites with small gold nanoparticles of 0.7 nm radius. We apply AXSI to two different cysteine-variants of maltose binding protein in the presence and absence of its ligand maltose and find distances in quantitative agreement with single-molecule FRET experiments. Our study shows that AXSI enables determination of intramolecular distance distributions under virtually arbitrary solution conditions and we anticipate its broad use to characterize protein conformational ensembles and dynamics.

• Dokumententyp: Zeitschriftenartikel
• EU Funded Grant Agreement Number: 101002656
• EU-Projekte: Horizon 2020 > ERC Grants ; Horizon 2020 > ERC Grants > ERC Consolidator Grant
• Fakultät: Physik ; Biologie
• Themengebiete: 500 Naturwissenschaften und Mathematik > 530 Physik ; 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften; Biologie
• ISSN: 2040-3364
• Bemerkung: First published 09 Dec 2024
• Sprache: Englisch
• DFG: Gefördert durch die Deutsche Forschungsgemeinschaft (DFG) - 111166240