Biology subjects

Gil-Garcia, M.

Publications and source records attributed to Gil-Garcia, M..

3 recordsLinked to original sources

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↗

bioRxiv2024

Enhancement of Enzymatic Activity by Biomolecular Condensates through pH Buffering

Biomolecular condensates can affect enzymatic reactions by locally changing not only concentrations of molecules but also their environment. Since protein conformations can differ between the dense and dilute phase, phase separation can particularly modulate enzymes characterized by a conformation-dependent activity. Here, we generate enzymatic condensates based on a lipase from Bacillus thermocatenulatus, which exhibits an equilibrium between a closed, inactive state, and an open, active conformation. We show that the activity of the enzyme increases inside the dense phase, leading to an enhancement of the overall reaction rate in the phase-separated system. Moreover, we demonstrate that these condensates can generate a more basic environment compared to the surrounding solution. As a consequence, the phase-separated system maintains a high enzymatic activity even in acidic conditions that would be otherwise less favourable for the lipase. We further show that the formation of two phases with distinct pH values optimizes a cascade reaction involving two enzymes with different optimal pH conditions. Our results demonstrate that biomolecular condensates can also affect the dependence of enzymatic rates on solution pH, thereby expanding the accessible pH interval and enabling network reactions with enzymes that require distinct pH values. These findings have crucial implications in biology and biotechnology for biocatalytic engineering.

biochemistry↗

bioRxiv2022

Cryo-EM Structure of a Mammalian-specific Alternative Amyloid Exon

hnRNPDL is a ribonucleoprotein (RNP) involved in transcription and RNA-processing, with missense mutations causing limb-girdle muscular dystrophy-3 (LGMDD3). Mammalian-specific alternative splicing (AS) renders three natural isoforms, hnRNPDL-2 being predominant in humans. We present the cryo-electron microscopy structure of full-length hnRNPDL-2 amyloid fibrils, which are stable, non-toxic, and bind nucleic acids, with the RNA binding domains building a solenoidal coat around them. The amyloid core consists of a single Gly/Tyr-rich and highly hydrophilic filament containing internal water channels. The architecture and activity of hnRNPDL-2 fibrils are reminiscent of functional amyloids, our results suggesting that LGMDD3 might be a loss-of-function disease associated with impaired fibrillation. Strikingly, the fibril core matches exon 6, absent in the soluble hnRNPDL-3 isoform. This provides structural evidence for AS controlling hnRNPDL assembly by precisely including/skipping an amyloid exon, a mechanism that holds the potential to generate functional diversity in RNPs.

biochemistry↗