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Nature segregates fundamental tasks such as information storage/transmission and catalysis between two different compound classes (e.g. polynucleotides for replication and folded polyamides for catalysis). This division of labor is likely a product of evolution, raising the question of how simpler systems in which replicators and folded macromolecules co‐exist may emerge in the transition from chemistry to biology. In synthetic systems, achieving co‐existence of replicators and foldamers in a single molecular network remains an unsolved problem. Previous work on dynamic molecular networks has given rise to either self‐replicating fibers or well‐defined foldamer structures (or completely un‐sorted complex systems). We report a system in which two cross‐reactive dithiol (nucleobase‐ and peptide‐based) building blocks self‐sort into a replicator fiber and foldamer that both emerge spontaneously and co‐exist. The self‐sorting behavior remains prevalent across different building block ratios as two phases of emergence occur: replicator growth followed by foldamer formation. This is attributed to the autocatalytic formation of the replicator fiber, followed by enrichment of the system in the remaining building block, which is subsequently incorporated into a foldamer., Two different dithiol building blocks interconvert through reversible covalent bonds to initially form a dynamic combinatorial library of macrocycles of various size. The self‐sorting process starts by the amplification of a single library member to form self‐replicating fibers. This step is followed by enrichment of the system in the remaining building block, which forms a large macrocyclic foldamer.

Unravelling how the complexity of living systems can (have) emerge(d) from simple chemical reactions is one of the grand challenges in contemporary science. Evolving systems of self‐replicating molecules may hold the key to this question. Here we show that, when a system of replicators is subjected to a regime where replication competes with replicator destruction, simple and fast replicators can give way to more complex and slower ones. The structurally more complex replicator was found to be functionally more proficient in the catalysis of a model reaction. These results show that chemical fueling can maintain systems of replicators out of equilibrium, populating more complex replicators that are otherwise not readily accessible. Such complexification represents an important requirement for achieving open‐ended evolution as it should allow improved and ultimately also new functions to emerge., Chemical fueling enables complex self‐replicating molecules to outcompete structurally simpler and faster replicators, revealing an important mechanism behind complexification that is required to transform chemistry into biology.

Unraveling how chemistry can give rise to biology is one of the greatest challenges of contemporary science. Achieving life-like properties in chemical systems is therefore a popular topic of research. Synthetic chemical systems are usually deterministic: the outcome is determined by the experimental conditions. In contrast, many phenomena that occur in nature are not deterministic but caused by random fluctuations (stochastic). Here, we report on how, from a mixture of two synthetic molecules, two different self-replicators emerge in a stochastic fashion. Under the same experimental conditions, the two self-replicators are formed in various ratios over several repeats of the experiment. We show that this variation is caused by a stochastic nucleation process and that this stochasticity is more pronounced close to a phase boundary. While stochastic nucleation processes are common in crystal growth and chiral symmetry breaking, it is unprecedented for systems of synthetic self-replicators.

Self-assembly features prominently in fields ranging from materials science to biophysical chemistry. Assembly pathways, often passing through transient intermediates, can control the outcome of assembly processes. Yet, the mechanisms of self-assembly remain largely obscure due to a lack of experimental tools for probing these pathways at the molecular level. Here, the self-assembly of self-replicators into fibers is visualized in real-time by high-speed atomic force microscopy (HS-AFM). Fiber growth requires the conversion of precursor molecules into six-membered macrocycles, which constitute the fibers. HS-AFM experiments, supported by molecular dynamics simulations, revealed that aggregates of precursor molecules accumulate at the sides of the fibers, which then diffuse to the fiber ends where growth takes place. This mechanism of precursor reservoir formation, followed by one-dimensional diffusion, which guides the precursor molecules to the sites of growth, reduces the entropic penalty associated with colocalizing precursors and growth sites and constitutes a new mechanism for supramolecular polymerization.

The conditions that led to the formation of the first organisms and the ways that life originates from a lifeless chemical soup are poorly understood. The recent hypothesis of ''RNA-peptide coevolution'' suggests that the current close relationship between amino acids and nucleobases may well have extended to the origin of life. We now show how the interplay between these compound classes can give rise to new self-replicating molecules using a dynamic combinatorial approach. We report two strategies for the fabrication of chimeric amino acid/nucleobase self-replicating macrocycles capable of exponential growth. The first one relies on mixing nucleobase- and peptide-based building blocks, where the ligation of these two gives rise to highly specific chimeric ring structures. The second one starts from peptide nucleic acid (PNA) building blocks in which nucleobases are already linked to amino acids from the start. While previously reported nucleic acid-based self-replicating systems rely on pre-synthesis of (short) oligonucleotide sequences, self-replication in the present systems start from units containing only a single nucleobase. Self-replication is accompanied by self-assembly, spontaneously giving rise to an ordered one-dimensional arrangement of nucleobase nanostructures.

The ability of molecules and systems to make copies of themselves and the ability of molecules to fold into stable, well-defined three-dimensional conformations are of considerable importance in the formation and persistence of life. The question of how, during the emergence of life, oligomerization reactions become selective and channel these reactions toward a small number of specific products remains largely unanswered. Herein, we demonstrate a fully synthetic chemical system where structurally complex foldamers and self-replicating assemblies emerge spontaneously and with high selectivity from pools of oligomers as a result of forming noncovalent interactions. Whether foldamers or replicators form depends on remarkably small differences in building block structures and composition and experimental conditions. We also observed the dynamic transformation of a foldamer into a replicator. These results show that the structural requirements/design criteria for building blocks that lead to foldamers are similar to those that lead to replicators. What determines whether folding or replication takes place is not necessarily the type of noncovalent interaction, but only whether they occur intra- or intermolecularly. This work brings together, for the first time, the fields of replicator and foldamer chemistry.

This dissertation is focused on two major investigations. The first one is about the mechanism with which a known self-replicator can create copies of itself. This study led to the first real-time visualization of a molecule capable of self-replication. The second is about the introduction of additional functionalities within self-replicating systems of molecules. Here, we were able to show emergent behavior: catalytic activity that would only start when the self-replication process takes place.

Ultra-performance liquid chromatography is a common analysis tool, and stirring is common in many laboratory setups. Here we show a device which enables continuous stirring of samples whilst inside an ultra-performance liquid chromatography system. Utilizing standard magnetic stirring bars that fit standard vials, the device allows for the automation of experimental setups that require stirring. The device is designed such that it can replace the standard sample holder and fits in its place, while being battery operated. The use of three-dimensional (3D) printing and commercially available parts enables low-effort and low-cost device production, as well as easy modifications. Testing the device was performed by video analysis and by following the kinetics of a dynamic combinatorial library that is known to be exquisitely sensitive to agitation, as a result of involving a fiber growth-breakage mechanism. Design files and schematics are provided. High- and ultra-performance liquid chromatography are valuable tools for the identification of components in complex mixtures, but these instruments lack sample stirring capabilities. Here the authors design an automated device that enables continuous stirring of samples inside an ultra-performance liquid chromatography system, and can be reproduced and modified using 3D printing technology.

A device is presented, which enables continuous stirring of samples whilst inside an ultra-performance liquid chromatography system. Utilizing standard magnetic stirring bars that fit standard vials, the device allows for the automation of experimental setups that require stirring. The device is designed such that it can replace the standard sample holder and fits in its place, while being battery operated. The use of 3D printing and commercially available parts enables low-effort and low-cost device production, as well as easy modifications. Various tests were performed by following the kinetics of a dynamic combinatorial library that is known for exhibiting self-replication under mechanical agitation, via fiber growth-breakage mechanism. Design files and schematics are available.

This work addressed how self-replicating molecules can evolve to become more complex. Subjecting a system in which two self-replicating molecules compete for a common food source to a regime in which they are both degraded leads to survival of the most complex of the two replicators, even though this replicator is less efficient at replicating. These results show that chemical fueling of a replication process can drive the complexification of the replicator. The more complex replicator is also more proficient at catalyzing a model reaction, showing that complexification can also enhance (catalytic) function.

In this manuscript we describe how, by taking a systems approach, complex energy landscapes of supramolecular assemblies can be navigated using stoichiometry to control, with remarkable selectivity, which assembly gets populated. The perhaps counterintuitive finding is that it is not necessarily the assembly that, in a one-to-one comparison, is the most stable that wins the competition for common building blocks, even though the system is under thermodynamic control. Instead, an individually less stable assembly may completely dominate the system. This domination is possible when the building block stoichiometry in the system matches the stoichiometry of this specific assembly, allowing the system to maximize binding energy by making a large number of assemblies of moderate stability as opposed to a small number of more stable assemblies.

How life can emerge from inanimate matter is one of the grand questions in science. Self-replicating molecules are necessary for the transition from chemistry to biology, but they need to acquire additional functions for life to emerge. Catalysis is one of the most essential of such functionalities, but mechanisms through which self-replicators can acquire catalytic and, in extension, metabolic properties have remained elusive. Here we show how catalytic activity and promiscuity in a self-replicator emerges through co-option: features that are selected to benefit replication inadvertently result in an arrangement of chemical functionalities that is conducive to catalysis. Specifically, we report self-assembly driven self-replicators that promote both a model retro-aldol reaction and the cleavage of fluorenylmethoxycarbonyl groups, with the latter transformation exerting a positive feedback on replication (protometabolism). Such chance invention of new function at the molecular level marks a pivotal step toward the de novo synthesis of life. In the development of chemical complexity—and the transition into biology—developing catalytic functionality is essential. Here the authors report the emergence of catalytic activity for two separate reactions (including one demonstrating a positive feedback on replication) in a self-replicating system.