Biophysical Society Thematic Meeting | Tutzing 2026

Single-Molecule FRET: The Next 30 Years

Poster Abstracts

18-POS Board 10 FAITHFUL TETHERING: DISORDERED LINKERS AS A TOOL TO STUDY INTERACTIONS OF CHARGED INTRINSICALLY DISORDERED PROTEINS

Lucia Franchini 1 ; Soundhararajan Gopi 1 ; Daniel Nettels 1 ; Ben Schuler 1 ; 1 University of Zurich, Department of Biochemistry, Zurich, Switzerland

The paradigm that the affinity of biomolecular interactions relies on well-defined structures is challenged by the complex formed between the highly and oppositely charged intrinsically disordered proteins linker histone H1.0 (H1) and prothymosin α (ProT α ). H1 and ProT α form a highly dynamic, disordered complex that lacks a structured binding interface and yet has very high affinity [1]. In this project, we use smFRET to investigate H1-ProT α interaction using engineered fusion constructs, in which H1 and ProT α are tethered by an uncharged, disordered peptide linker. By varying the linker length we tune the effective local concentration of the binding partners, and dissect how the linkers affect H1-ProT α interactions. Our results show that the linker minimally perturbs the conformational ensemble, dynamics, oligomerization and extreme ionic strength sensitivity of the native H1-ProT α interaction. Coarse-grained molecular dynamics simulations reproduce and illustrate these experimental trends. The fusion constructs now allow us to investigate more complex interactions enabled by highly disordered complexes, such as the very short-lived ternary complexes that can lead to very rapid binding kinetics despite their high affinity [2]. A notable example is the ProT α -driven displacement of H1 from the nucleosome [3] by means of a ternary complex formed by ProT α , H1 and the nucleosome. The tethered H1-ProT α fusion construct allows us to populate this ternary complex at equilibrium and study its properties directly. Moreover, the fusion construct may allow us to probe the properties of the ProT α -H1 interactions in live cells.[1] Borgia, A., Borgia, M., Bugge, K. et al., Nature, 555, 61–66 (2018).[2] Sottini, A., Borgia, A., Borgia, M.B. et al., Nature Communications, 11, 5736 (2020).[3] Heidarsson, P.O., Mercadante, D., Sottini, A. et al., Nature Chemistry, 14, 224–231 (2022).

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