Sequential Design of Experiments enables accessible and efficient multidimensional mapping of single- and multi-protein condensates
- 1Biozentrum, University of Basel, Basel, Switzerland
Abstract
Biomolecular condensation emerges from networks of multivalent interactions that respond collectively to molecular composition and environmental conditions. Here, we establish a sequential Design of Experiments (DoE) framework to systematically and efficiently characterise multidimensional condensation landscapes. Applied to three RNA-binding condensate proteins - Dhh1, Ded1 and PGL-3 - the workflow revealed unexpected protein-specific interactions between biophysical and biochemical variables, including RNA, pH and ionic strength, and uncovered context-dependent switch-like behaviours. Extending the framework to protein mixtures revealed how proteins affected each other's condensation and whether they formed condensate-within-condensate structures under different experimental conditions. Finally, an unexpected response identified by DoE prompted mechanistic analysis of Dhh1, revealing an ATP-independent RNA-binding surface that promotes efficient condensation in vitro and contributes to P-body formation in cells. Sequential DoE therefore provides an experimentally accessible strategy for uncovering context-dependent condensate regulation and converting multidimensional response patterns into mechanistic hypotheses.
Related
- MOSHPIT: Accessible, reproducible metagenome data science on the QIIME 2 framework
- Long-range transport of biomolecular condensates by Marangoni effect
- Transmembrane coupling of protein condensates via membrane-mediated interactions: A simulation study
- To mix or to de-mix?: Cation-π suppresses heterogeneity in local structure and dynamics of heterotypic Condensates
- Active RNA synthesis patterns nuclear condensates
BibTeX
@misc{siketanc2026sequential,
title = {{Sequential Design of Experiments enables accessible and efficient multidimensional mapping of single- and multi-protein condensates}},
author = {M. Siketanc and F. Weidner and C. M. Nuesperling and M. Hondele},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.29.755056},
url = {https://www.biorxiv.org/content/10.64898/2026.09.29.755056v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Sequential Design of Experiments enables accessible and efficient multidimensional mapping of single- and multi-protein condensates
AU - M. Siketanc
AU - F. Weidner
AU - C. M. Nuesperling
AU - M. Hondele
PY - 2026
DA - 2026/09/29
PB - bioRxiv
DO - 10.64898/2026.09.29.755056
UR - https://www.biorxiv.org/content/10.64898/2026.09.29.755056v1
AB - Biomolecular condensation emerges from networks of multivalent interactions that respond collectively to molecular composition and environmental conditions. Here, we establish a sequential Design of Experiments (DoE) framework to systematically and efficiently characterise multidimensional condensation landscapes. Applied to three RNA-binding condensate proteins - Dhh1, Ded1 and PGL-3 - the workflow revealed unexpected protein-specific interactions between biophysical and biochemical variables, including RNA, pH and ionic strength, and uncovered context-dependent switch-like behaviours. Extending the framework to protein mixtures revealed how proteins affected each other's condensation and whether they formed condensate-within-condensate structures under different experimental conditions. Finally, an unexpected response identified by DoE prompted mechanistic analysis of Dhh1, revealing an ATP-independent RNA-binding surface that promotes efficient condensation in vitro and contributes to P-body formation in cells. Sequential DoE therefore provides an experimentally accessible strategy for uncovering context-dependent condensate regulation and converting multidimensional response patterns into mechanistic hypotheses.
ER -