Biology subjects

Buehler, M. A.

Publications and source records attributed to Buehler, M. A..

2 recordsLinked to original sources

bioRxiv2026

Engineering nanocondensate formation through sequence composition and patterning

Many biological proteins can assemble into dynamic, non-stoichiometric structures known as biomolecular condensates. Although typically observed at micrometer scales in vitro, recent evidence shows that these condensates often appear as nanoscale assemblies both in vitro and in cells. Moreover, biochemical reactions can be more efficiently promoted in nanoscale condensates than in micron-sized droplets, due to mass-transfer limitations and interfacial effects. Therefore, in analogy with colloids, the function of condensate materials can be engineered by tuning their size distribution. However, controlling the size of condensates remains challenging, as the molecular mechanisms that prevent small condensates from coarsening into larger ones are still poorly understood. Here, we developed and applied a computational pipeline that combines high-throughput molecular simulations, machine learning, and mixed-integer linear programming to design phase-separating peptides that form metastable nanocondensates across a broad range of experimental conditions. In addition to experimentally validating these peptides, we elucidate the underlying molecular mechanisms and derive initial design rules. In particular, we show that scaffold net charge combined with sequence blockiness can lead to high phase separation propensity while simultaneously yielding low interfacial tension, thereby slowing ripening. Moreover, these combined properties induce an electrostatics-driven alignment of molecules at the interface, which generates an additional size-dependent coalescence barrier. We further show that these features are shared by biological proteins, providing a possible mechanistic basis for the widespread occurrence of nanocondensates in biological systems. Altogether, our findings shed light on the molecular mechanisms behind nanocondensate formation, and provide a platform to design nanocondensates for several potential applications in bioengineering and biotechnology. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/706365v2_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@17c379aorg.highwire.dtl.DTLVardef@26686dorg.highwire.dtl.DTLVardef@19206a7org.highwire.dtl.DTLVardef@1f2da71_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2025

De novo design of peptides localizing at the interface of biomolecular condensates

The interface of biomolecular condensates has been shown to play an important role in processes such as protein aggregation and biochemical reactions. Targeted modulation of these interfaces could, therefore, serve as an effective strategy for engineering condensates and modifying aberrant behaviors. However, the molecular grammar driving the preferential localization of molecules at condensate interfaces remains largely unknown. In this study, we developed a computational pipeline that combines highthroughput coarse-grained simulations, machine learning, and mixed-integer linear programming to design peptides that selectively partition at the interfaces of specific condensate targets. Using this workflow, we designed and synthesized peptides that localize at the interface of three distinct condensates formed by different intrinsically disordered protein regions (IDRs). These peptides exhibit surfactant-like architectures, with one tail incorporated into the condensate and the other excluded from the dense phase. In all cases, the tail entering the condensates is enriched in aromatic residues, while the sequence of the excluded tail varies among the IDRs. For hnRNPA1- and LAF1-IDRs, the excluded tail is enriched in lysines and matches the net charge of the condensate-forming protein, promoting electrostatic repulsion. In the case of DDX4-IDR, which exhibits the lowest charge density, the excluded tail mainly consists of uncharged valine residues, which exhibit negligible interactions with the scaffold protein. These results highlight the importance of the net charge of the scaffold as a key physicochemical parameter for designing peptides with preferential interfacial localization. Overall, our pipeline represents a promising strategy for the rational design of interface-localizing peptides and the identification of the corresponding molecular grammar. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=73 SRC="FIGDIR/small/653111v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@eab174org.highwire.dtl.DTLVardef@2852d6org.highwire.dtl.DTLVardef@156960corg.highwire.dtl.DTLVardef@1952e1d_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗