Native Mass Spectrometry Captures the Conformational Plasticity of Proteins with Low-Complexity Domains

Abstract

Disordered regions are an important functional feature of many multidomain proteins. A prime example is proteins in membraneless organelles, which contain folded domains that engage in specific interactions and disordered low-complexity (LC) domains that mediate liquid-liquid phase separation. Studying these complex architectures remains challenging due to their conformational variability. Native mass spectrometry (nMS) is routinely employed to analyze conformations and interactions of folded or disordered proteins; however, its ability to analyze proteins with disordered LC domains has not been investigated. Here, we analyze the ionization and conformational states of designed model proteins that recapitulate key features of proteins found in membraneless organelles. Our results show that charge state distributions (CSDs) in nMS reflect partial disorder regardless of the protein sequence, providing insights into their conformational plasticity and interactions. By applying the same CSD analysis to a spider silk protein fragment, we find that interactions between folded domains that trigger silk assembly simultaneously induce conformational changes in the LC domains. Lastly, using intact nucleosomes, we demonstrate that CSDs are a good predictor for the disorder content of complex native assemblies. We conclude that nMS reliably informs about the conformational landscape of proteins with LC domains, which is crucial for understanding protein condensates in cellular environments.

Keywords

intrinsic disorder; electrospray ionization; protein engineering; liquid-liquid phase separation

Published in

JACS Au
2025
Publisher: AMER CHEMICAL SOC

SLU Authors

• Rising, Anna

  • Department of Animal Biosciences, Swedish University of Agricultural Sciences
    
  • Karolinska Institutet

UKÄ Subject classification

Biochemistry and Molecular Biology

Publication identifier

• DOI: https://doi.org/10.1021/jacsau.4c00961

Permanent link to this page (URI)

https://res.slu.se/id/publ/140186