Polyubiquitin ligand-induced phase transitions are optimized by spacing between ubiquitin units.

Biomolecular condensates form via multivalent interactions among key macromolecules and are regulated through ligand binding and/or posttranslational modifications. One such modification is ubiquitination, the covalent addition of ubiquitin (Ub) or polyubiquitin chains to target macromolecules. Specific interactions between polyubiquitin chains and partner proteins, including hHR23B, NEMO, and UBQLN2, regulate condensate assembly or disassembly. Here, we used a library of designed polyubiquitin hubs and UBQLN2 as model systems for determining the driving forces of ligand-mediated phase transitions. Perturbations to either the UBQLN2-binding surface of Ub or the spacing between Ub units reduce the ability of hubs to modulate UBQLN2 phase behavior. By developing an analytical model based on polyphasic linkage principles that accurately described the effects of different hubs on UBQLN2 phase separation, we determined that introduction of Ub to UBQLN2 condensates incurs a significant inclusion energetic penalty. This penalty antagonizes the ability of polyUb hubs to scaffold multiple UBQLN2 molecules and cooperatively amplify phase separation. The extent to which polyubiquitin hubs promote UBQLN2 phase separation is encoded in the spacings between Ub units. This spacing is modulated by chains of different linkages and designed chains of different architectures, thus illustrating how the ubiquitin code regulates functionality via the emergent properties of the condensate. The spacing in naturally occurring linear polyubiquitin chains is already optimized to promote phase separation with UBQLN2. We expect our findings to extend to other condensates, emphasizing the importance of ligand properties, including concentration, valency, affinity, and spacing between binding sites in studies and designs of condensates.

Decoding optimal ligand design for multicomponent condensates.

Biomolecular condensates form via multivalent interactions among key macromolecules and are regulated through ligand binding and/or post-translational modifications. One such modification is ubiquitination, the covalent addition of ubiquitin (Ub) or polyubiquitin chains to target macromolecules for various cellular processes. Specific interactions between polyubiquitin chains and partner proteins, including hHR23B, NEMO, and UBQLN2, regulate condensate assembly or disassembly. Here, we used a library of designed polyubiquitin hubs and UBQLN2 as model systems for determining the driving forces of ligand-mediated phase transitions. Perturbations to the UBQLN2-binding surface of Ub or deviations from the optimal spacing between Ub units reduce the ability of hubs to modulate UBQLN2 phase behavior. By developing an analytical model that accurately described the effects of different hubs on UBQLN2 phase diagrams, we determined that introduction of Ub to UBQLN2 condensates incurs a significant inclusion energetic penalty. This penalty antagonizes the ability of polyUb hubs to scaffold multiple UBQLN2 molecules and cooperatively amplify phase separation. Importantly, the extent to which polyubiquitin hubs can promote UBQLN2 phase separation are encoded in the spacings between Ub units as found for naturally-occurring chains of different linkages and designed chains of different architectures, thus illustrating how the ubiquitin code regulates functionality via the emergent properties of the condensate. We expect our findings to extend to other condensates necessitating the consideration of ligand properties, including concentration, valency, affinity, and spacing between binding sites in studies and designs of condensates.

Synergic effect of aqueous extracts of Ocimum sanctum and Trigonella foenum-graecum L on the in situ green synthesis of silver nanoparticles and as a preventative agent against antibiotic-resistant food spoiling organisms.

The combination of Ocimum sanctum and Trigonella foenum-graecum L leaf water extract synergistically acts as a reducing and capping agent for the synthesis of narrow polydisperse silver nanoparticles (Ag NPs) with controlled sizes depending on the precursor (AgNO3) concentration in the plant extract. The toxicity of 40 nm-sized green synthesized Ag NPs is less than that of 10 nm-sized NPs. The Ag NP solution in Ocimum sanctum and Trigonella foenum-graecum L leaf water extract shows synergic antibacterial effect on Gram-negative bacteria by effecting the ester group of the lipids (hydrolysis) and also breaking the amide bonds of the bacterial chemical constituents, which leads to their rapid death.