A Carbodiimide-Fueled Reaction Cycle That Forms Transient 5(4H)-Oxazolones.

In life, molecular architectures, like the cytoskeletal proteins or the nucleolus, catalyze the conversion of chemical fuels to perform their functions. For example, tubulin catalyzes the hydrolysis of GTP to form a dynamic cytoskeletal network. In contrast, myosin uses the energy obtained by catalyzing the hydrolysis of ATP to exert forces. Artificial examples of such beautiful architectures are scarce partly because synthetic chemically fueled reaction cycles are relatively rare. Here, we introduce a new chemical reaction cycle driven by the hydration of a carbodiimide. Unlike other carbodiimide-fueled reaction cycles, the proposed cycle forms a transient 5(4H)-oxazolone. The reaction cycle is efficient in forming the transient product and is robust to operate under a wide range of fuel inputs, pH, and temperatures. The versatility of the precursors is vast, and we demonstrate several molecular designs that yield chemically fueled droplets, fibers, and crystals. We anticipate that the reaction cycle can offer a range of other assemblies and, due to its versatility, can also be incorporated into molecular motors and machines.

Carbodiimide-fueled catalytic reaction cycles to regulate supramolecular processes.

Using molecular self-assembly, supramolecular chemists can create Gigadalton-structures with angstrom precision held together by non-covalent interactions. However, despite relying on the same molecular toolbox for self-assembly, these synthetic structures lack the complexity and sophistication of biological assemblies. Those assemblies are non-equilibrium structures that rely on the constant consumption of energy transduced from the hydrolysis of chemical fuels like ATP and GTP, which endows them with dynamic properties, e.g., temporal and spatial control and self-healing ability. Thus, to synthesize life-like materials, we have to find a reaction cycle that converts chemical energy to regulate self-assembly. We and others recently found that this can be done by a reaction cycle that hydrates carbodiimides. This feature article aims to provide an overview of how the energy transduced from carbodiimide hydration can alter the function of molecules and regulate molecular assemblies. The goal is to offer the reader design considerations for carbodiimide-driven reaction cycles to create a desired morphology or function of the assembly and ultimately to push chemically fueled self-assembly further towards the bottom-up synthesis of life.

Droplet Formation by Chemically Fueled Self-Assembly: The Role of Precursor Hydrophobicity.

We investigate active droplets that form at the expense of a chemical fuel in aqueous buffer and vanish autonomously. Dynamic light scattering reveals the scattered intensity, the hydrodynamic radius, and the width of the size distribution with high precision as well as high temporal and spatial resolutions. Comparing the resulting time-dependent behavior of the droplet characteristics with the time-dependent concentration of the anhydrides, the roles of the chemical reaction cycle and of colloidal growth processes are elucidated. The droplet sizes and lifetimes depend strongly on the hydrophobicity of the precursor, and the growth rate is found to correlate with the deactivation rate of the product.

Morphological transitions in chemically fueled self-assembly.

In chemically fueled self-assembly, a reaction cycle activates and deactivates molecules for self-assembly. The resulting assembly is dynamic and should be endowed with unique behavior in this kinetically controlled regime. Recent works have mainly focused on design rules for the activation of molecules for self-assembly, thereby assuming that disassembly upon deactivation inherently follows. However, that is not always the case. This work shows a family of peptides that assemble into colloids regulated through a chemical reaction cycle. Despite their similarity in assembly, we find that they follow a different disassembly pathway upon deactivation. The colloids from several peptides completely disassemble as fuel depletes while others transition into fibers. Our findings demonstrate that assembly and disassembly should be taken into account in chemically fueled self-assembly.

Active coacervate droplets as a model for membraneless organelles and protocells.

Membraneless organelles like stress granules are active liquid-liquid phase-separated droplets that are involved in many intracellular processes. Their active and dynamic behavior is often regulated by ATP-dependent reactions. However, how exactly membraneless organelles control their dynamic composition remains poorly understood. Herein, we present a model for membraneless organelles based on RNA-containing active coacervate droplets regulated by a fuel-driven reaction cycle. These droplets emerge when fuel is present, but decay without. Moreover, we find these droplets can transiently up-concentrate functional RNA which remains in its active folded state inside the droplets. Finally, we show that in their pathway towards decay, these droplets break apart in multiple droplet fragments. Emergence, decay, rapid exchange of building blocks, and functionality are all hallmarks of membrane-less organelles, and we believe that our work could be powerful as a model to study such organelles.

Self-assembled multivalent (SAMul) ligand systems with enhanced stability in the presence of human serum.

Self-assembled cationic micelles are an attractive platform for binding biologically-relevant polyanions such as heparin. This has potential applications in coagulation control, where a synthetic heparin rescue agent could be a useful replacement for protamine, which is in current clinical use. However, micelles can have low stability in human serum and unacceptable toxicity profiles. This paper reports the optimisation of self-assembled multivalent (SAMul) arrays of amphiphilic ligands to bind heparin in competitive conditions. Specifically, modification of the hydrophobic unit kinetically stabilises the self-assembled nanostructures, preventing loss of binding ability in the presence of human serum - cholesterol hydrophobic units significantly outperform systems with a simple aliphatic chain. It is demonstrated that serum albumin disrupts the binding thermodynamics of the latter system. Molecular simulation shows aliphatic lipids can more easily be removed from the self-assembled nanostructures than the cholesterol analogues. This agrees with the experimental observation that the cholesterol-based systems undergo slower disassembly and subsequent degradation via ester hydrolysis. Furthermore, by stabilising the SAMul nanostructures, toxicity towards human cells is decreased and biocompatibility enhanced, with markedly improved survival of human hepatoblastoma cells in an MTT assay.

Dissipative Self-Assembly of Photoluminescent Silicon Nanocrystals.

Solutions of silicon nanocrystals (SiNCs) are used in a diverse range of applications because of their tunable photoluminescence, biocompatibility, and the abundance of Si. In dissipative supramolecular materials, self-assembly of molecules or nanoparticles is driven by a chemical reaction network that irreversible consumes fuel. The properties of the emerging structures are controlled by the kinetics of the underlying chemical reaction network. Herein, we demonstrate the dissipative self-assembly of photoluminescent SiNCs driven by a chemical fuel. A chemical reaction induces self-assembly of the water-soluble SiNCs. However, the assemblies are transient, and when the chemical reaction network runs out of fuel, the SiNCs disassemble. The lifetime of the assemblies is controlled by the amount of fuel added. As an application of the transient supramolecular material, we demonstrate that the platform can be used to control the delayed uptake of the nanocrystals by mammalian cells.

Self-selection of dissipative assemblies driven by primitive chemical reaction networks.

Life is a dissipative nonequilibrium structure that requires constant consumption of energy to sustain itself. How such an unstable state could have selected from an abiotic pool of molecules remains a mystery. Here we show that liquid phase-separation offers a mechanism for the selection of dissipative products from a library of reacting molecules. We bring a set of primitive carboxylic acids out-of-equilibrium by addition of high-energy condensing agents. The resulting anhydrides are transiently present before deactivation via hydrolysis. We find the anhydrides that phase-separate into droplets to protect themselves from hydrolysis and to be more persistent than non-assembling ones. Thus, after several starvation-refueling cycles, the library self-selects the phase-separating anhydrides. We observe that the self-selection mechanism is more effective when the library is brought out-of-equilibrium by periodic addition of batches as opposed to feeding it continuously. Our results suggest that phase-separation offers a selection mechanism for energy dissipating assemblies.

Dissipative assemblies that inhibit their deactivation.

Dissipative self-assembly is a process in which energy-consuming chemical reaction networks drive the assembly of molecules. Prominent examples from biology include the GTP-fueled microtubule and ATP-driven actin assembly. Pattern formation and oscillatory behavior are some of the unique properties of the emerging assemblies. While artificial counterparts exist, researchers have not observed such complex responses. One reason for the missing complexity is the lack of feedback mechanisms of the assemblies on their chemical reaction network. In this work, we describe the dissipative self-assembly of colloids that protect the hydrolysis of their building blocks. The mechanism of inhibition is generalized and explored for other building blocks. We show that we can tune the level of inhibition by the assemblies. Finally, we show that the robustness of the assemblies towards starvation is affected by the degree of inhibition.

Non-equilibrium dissipative supramolecular materials with a tunable lifetime.

Many biological materials exist in non-equilibrium states driven by the irreversible consumption of high-energy molecules like ATP or GTP. These energy-dissipating structures are governed by kinetics and are thus endowed with unique properties including spatiotemporal control over their presence. Here we show man-made equivalents of materials driven by the consumption of high-energy molecules and explore their unique properties. A chemical reaction network converts dicarboxylates into metastable anhydrides driven by the irreversible consumption of carbodiimide fuels. The anhydrides hydrolyse rapidly to the original dicarboxylates and are designed to assemble into hydrophobic colloids, hydrogels or inks. The spatiotemporal control over the formation and degradation of materials allows for the development of colloids that release hydrophobic contents in a predictable fashion, temporary self-erasing inks and transient hydrogels. Moreover, we show that each material can be re-used for several cycles.

Dissipative out-of-equilibrium assembly of man-made supramolecular materials.

The use of dissipative self-assembly driven by chemical reaction networks for the creation of unique structures is gaining in popularity. In dissipative self-assembly, precursors are converted into self-assembling building blocks by the conversion of a source of energy, typically a photon or a fuel molecule. The self-assembling building block is intrinsically unstable and spontaneously reverts to its original precursor, thus giving the building block a limited lifetime. As a result, its presence is kinetically controlled, which gives the associated supramolecular material unique properties. For instance, formation and properties of these materials can be controlled over space and time by the kinetics of the coupled reaction network, they are autonomously self-healing and they are highly adaptive to small changes in their environment. By means of an example of a biological dissipative self-assembled material, the unique concepts at the basis of these supramolecular materials will be discussed. We then review recent efforts towards man-made dissipative assembly of structures and how their unique material properties have been characterized. In order to help further the field, we close with loosely defined design rules that are at the basis of the discussed examples.

Emergent Catalytic Behavior of Self-Assembled Low Molecular Weight Peptide-Based Aggregates and Hydrogels.

We report a series of short peptides possessing the sequence (FE)n or (EF)n and bearing l-proline at their N-terminus that self-assemble into high aspect ratio aggregates and hydrogels. We show that these aggregates are able to catalyze the aldol reaction, whereas non-aggregated analogues are catalytically inactive. We have undertaken an analysis of the results, considering the accessibility of catalytic sites, pKa value shifts, and the presence of hydrophobic pockets. We conclude that the presence of hydrophobic regions is indeed relevant for substrate solubilization, but that the active site accessibility is the key factor for the observed differences in reaction rates. The results presented here provide an example of the emergence of a new chemical property caused by self-assembly, and support the relevant role played by self-assembled peptides in prebiotic scenarios. In this sense, the reported systems can be seen as primitive aldolase I mimics, and have been successfully tested for the synthesis of simple carbohydrate precursors.

Tetrapeptidic molecular hydrogels: self-assembly and co-aggregation with amyloid fragment Aß1-40.

A new family of isomeric tetrapeptides containing aromatic and polar amino acid residues that are able to form molecular hydrogels following a smooth change in pH is described. The hydrogels have been studied by spectroscopic and microscopic techniques showing that the peptide primary sequence has an enormous influence on the self-assembly process. In particular, the formation of extended hydrophobic regions and the appearance of π-stacking interactions have been revealed as the driving forces for aggregation. Moreover, the interaction of these compounds with amyloid peptidic fragment Aß1-40 has been studied and some of them have been shown to act as templates for the aggregation of this peptide into non-ß-sheet fibrillar structures. These compounds could potentially be used for the capture of toxic, soluble amyloid oligomers.