Self-replicating Molecules Research Articles

Can Chirality Answer Whether We Are Alone?

Detecting biosignatures of life in extraterrestrial environments remains one of the primary objectives of scientific inquiry. Currently, both remote and direct detection methods are primarily aimed at identifying key molecular classes fundamental to terrestrial biology. However, a more universally applicable spectroscopic approach could involve searching for homochiral molecules. Thus, this perspective delves into the significance of homochirality as a critical factor in the origin of life. Without homochirality, the formation of self-recognizing and self-replicating complex molecules would be hindered. The various hypotheses concerning the origin of homochiral molecules have been explored and analyzed within this context. This perspective emphasizes the potential for discovering extraterrestrial microscopic life through the detection of homochiral molecules using chirodetecting methods such as chromatography and chiroptical spectroscopy or circular polarimetry as a promising remote technique. This discussion highlights the importance of homochirality in the broader search for life beyond Earth and underscores the need for innovative methodologies and instrumentation in astrobiological research. These techniques can be an effective method for detecting homochirality on future planetary missions.

New Generation Self-Replicating RNA Vaccines Derived from Pestivirus Genome.

While mRNA vaccines have shown their worth, they have the same failing as inactivated vaccines, namely they have limited half-life, are non-replicating, and therefore limited to the size of the vaccine payload for the amount of material translated. New advances averting these problems are combining replicon RNA (RepRNA) technology with nanotechnology. RepRNA are large self-replicating RNA molecules (typically 12-15kb) derived from viral genomes defective in at least one essential structural protein gene. They provide sustained antigen production, effectively increasing vaccine antigen payloads over time, without the risk of producing infectious progeny. The major limitations with RepRNA are RNase-sensitivity and inefficient uptake by dendritic cells (DCs), which need to be overcome for efficacious RNA-based vaccine design. We employed biodegradable delivery vehicles to protect the RepRNA and promote DC delivery. Condensing RepRNA with polyethylenimine (PEI) and encapsulating RepRNA into novel Coatsome-replicon vehicles are two approaches that have proven effective for delivery to DCs and induction of immune responses in vivo.

Enantioselective Self-Replicators.

Self-replicating molecules provide a simple approach for investigating fundamental processes in scenarios of the emergence of life. Although homochirality is an important aspect of life and of how it emerged, the effects of chirality on self-replicators have received only little attention so far. Here, we report several self-assembled self-replicators with enantioselectivity that emerge spontaneously and grow only from enantiopure material. These require a relatively small number of chiral units in the replicators (down to eight) and in the precursors (down to a single chiral unit), compared to the only other enantioselective replicator reported previously. One replicator was found to incorporate material of its own handedness with high fidelity when provided with a racemic mixture of precursors, thus sorting (L)- and (D)-precursors into (L)- and (D)-replicators. Systematic studies reveal that the presence or absence of enantioselectivity depends on structural features (ring size of the replicator) that appear to impose constraints on its supramolecular organization. This work reveals new aspects of the little researched interplay between chirality and self-replication and represents another step toward the de novo synthesis of life.

Loading of Amino Acids onto RNA in a Putative RNA-Peptide World.

RNA is a molecule that can both store genetic information and perform catalytic reactions. This observed dualism places RNA into the limelight of concepts about the origin of life. The RNA world concept argues that life started from self-replicating RNA molecules, which evolved toward increasingly complex structures. Recently, we demonstrated that RNA, with the help of conserved non-canonical nucleosides, which are also putative relics of an early RNA world, had the ability to grow peptides covalently connected to RNA nucleobases, creating RNA-peptide chimeras. It is conceivable that such molecules, which combined the information-coding properties of RNA with the catalytic potential of amino acid side chains, were once the structures from which life emerged. Herein, we report prebiotic chemistry that enabled the loading of both nucleosides and RNAs with amino acids as the first step toward RNA-based peptide synthesis in a putative RNA-peptide world.

Did disordered proteins help launch life on Earth?

Four and a half billion years ago, our planet was a jagged, rocky landscape rich in minerals, swirling with carbon dioxide and other gases, and bombarded with meteorites. Somehow, around this time, the chemistry at work in this harsh environment yielded molecules that started to self-replicate. Thus began life on Earth. Most researchers studying early life believe that the first self-replicating molecule to take hold was RNA, an idea known as the RNA-world hypothesis. But eventually came proteins—the actors that carry out almost all cellular functions—and DNA, the blueprint that encodes them. It’s the proteins—and their building blocks, amino acids—that have always intrigued biochemist A. Keith Dunker, an emeritus professor at Indiana University. He knew that researchers had long speculated that the early genetic code was much simpler than it is today, perhaps just 2 or 3 DNA nucleotides coding for 12 or 14 amino acids rather than today’s canonical

Emergence of light-driven protometabolism on recruitment of a photocatalytic cofactor by a self-replicator.

Establishing how life can emerge from inanimate matter is among the grand challenges of contemporary science. Chemical systems that capture life's essential characteristics-replication, metabolism and compartmentalization-offer a route to understanding this momentous process. The synthesis of life, whether based on canonical biomolecules or fully synthetic molecules, requires the functional integration of these three characteristics. Here we show how a system of fully synthetic self-replicating molecules, on recruiting a cofactor, acquires the ability to transform thiols in its environment into disulfide precursors from which the molecules can replicate. The binding of replicator and cofactor enhances the activity of the latter in oxidizing thiols into disulfides through photoredox catalysis and thereby accelerates replication by increasing the availability of the disulfide precursors. This positive feedback marks the emergence of light-driven protometabolism in a system that bears no resemblance to canonical biochemistry and constitutes a major step towards the highly challenging aim of creating a new and completely synthetic form of life.

Spontaneous Emergence of Self-Replicating Molecules Containing Nucleobases and Amino Acids.

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 presynthesis 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.

Survival of Self-Replicating Molecules under Transient Compartmentalization with Natural Selection.

The problem of the emergence and survival of self-replicating molecules in origin-of-life scenarios is plagued by the error catastrophe, which is usually escaped by considering effects of compartmentalization, as in the stochastic corrector model. By addressing the problem in a simple system composed of a self-replicating molecule (a replicase) and a parasite molecule that needs the replicase for copying itself, we show that transient (rather than permanent) compartmentalization is sufficient to the task. We also exhibit a regime in which the concentrations of the two kinds of molecules undergo sustained oscillations. Our model should be relevant not only for origin-of-life scenarios but also for describing directed evolution experiments, which increasingly rely on transient compartmentalization with pooling and natural selection.

Design of conditions for self-replication.

A "self-replicator" is usually understood to be an object of definite form that promotes the conversion of materials in its environment into a nearly identical copy of itself. The challenge of engineering novel, micro- or nanoscale self-replicators has attracted keen interest in recent years, both because exponential amplification is an attractive method for generating high yields of specific products and, also, because self-reproducing entities have the potential to be optimized or adapted through rounds of iterative selection. Substantial steps forward have been achieved both in the engineering of particular self-replicating molecules and in the characterization of the physical basis for possible mechanisms of self-replication. At present, however, there is a need for a theoretical treatment of what physical conditions are most conducive to the emergence of novel self-replicating structures from a reservoir of building blocks on a desired time scale. Here we report progress in addressing this need. By analyzing the kinetics of a toy chemical model, we demonstrate that the emergence of self-replication can be controlled by coarse, tunable features of the chemical system, such as the fraction of fast reactions and the width of the rate constant distribution. We also find that the typical mechanism is dominated by the cooperation of multiple interconnected reaction cycles as opposed to a single isolated cycle. The quantitative treatment presented here may prove useful for designing novel self-replicating chemical systems.

Extreme and Topological Dissipative Solitons with Structured Matter and Structured Light.

Structuring of matter with nanoobjects allows one to generate soliton-like light bundles with extreme characteristics—temporal duration and spatial dimensions. On the other hand, structuring of light gives the possibility to form light bundles with complicated internal structure; their topology could be used for information coding similar to that in self-replicating RNA molecules carrying genetic code. Here we review the both variants of structuring. In the first variant, we consider a linear molecular chain and organic film interacting resonantly with laser radiation. Demonstrated are optical bistability, switching waves, and dissipative solitons, whose sizes for molecular J-aggregates can reach the nanometer range. We also discuss some theoretical approaches to take into account multi-particle interaction and correlations between molecules. In the second variant, light structuring in large-size laser medium with saturable amplification and absorption is achieved by preparation of the initial field distribution with a number of closed and unclosed vortex lines where the field vanishes. Various types of topological solitons, parameter domains of their stability, and transformation of the solitons with slow variation of the scheme parameters are presented.

Sijbren Otto

Angewandte Chemie International EditionVolume 57, Issue 33 p. 10416-10416 Author ProfileFree Access Sijbren Otto First published: 06 July 2018 https://doi.org/10.1002/anie.201806450AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Graphical Abstract The most important thing I learned from my parents is that when you do something, you do it properly. The biggest problem that scientists face is the metrics by which success is measured …” Find out more about Sijbren Otto in his Author Profile. Sijbren Otto The author presented on this page has published more than 10 articles since 2007 in Angewandte Chemie. His most recent contribution is published in this issue: “Parasitic Behavior of Self-Replicating Molecules ”: M. Altay, Y. Altay, S. Otto, Angew. Chem. Int. Ed. 2018, 57, 10564; Angew. Chem. 2018, 130, 10724. Date of birth: August 3, 1971 Position: Professor of Systems Chemistry, University of Groningen E-mail: s.otto@rug.nl Homepage: www.otto-lab.com ORCID: 0000-0003-0259-5637 Education: 1994 MSc, University of Groningen 1998 PhD with Prof. Dr. Jan B. F. N. Engberts, University of Groningen 1998 Postdoc with Prof. Dr. Steven L. Regen, Lehigh University 1999–2001 Marie Curie Fellow with Prof. Dr. Jeremy K. M. Sanders, University of Cambridge Awards: ERC Starting (2011) and Advanced (2016) Grants Current research interests: Systems chemistry; de novo synthesis of life; dynamic combinatorial chemistry; supramolecular chemistry; catalysis; origins of life Hobbies: Fossil hunting, hiking, korfball The most important thing I learned from my parents is that when you do something, you do it properly. The biggest problem that scientists face is the metrics by which success is measured. My favorite quote is “When you talk, you are only repeating what you already know. But if you listen, you may learn something new” (Dalai Lama XIV). I chose chemistry as a career because I thought it would provide a better chance for a job than what I ideally would have liked to become: a paleontologist. Chemistry was a good second choice though, as, unlike paleontology, it allows you to create something new. My most exciting discovery to date has been that working with self-replicating molecules allows combining the fun of chemistry with my fascination for evolution. My biggest motivation is to see if we can synthesize life in the lab, not by “simply” reconstructing a cell from its complex biological components, but by enabling it to emerge step by step from simple synthetic building blocks. My best investment was the five days I spent writing an ERC starting grant application (I do not normally earn € 30 000 per hour). Things happened this way because I only found out that, because of paternity leave, I was still eligible for it five days before the deadline. I did have to promise my wife not to make such investments again though. What I look for first in a publication is conceptual novelty. Amazing how few papers you really need to read once you take conceptual novelty as a filter. When I'm frustrated, I go running. Fortunately, the reverse is not true most of the time. I lose track of time when on a 12-hour flight back from Japan, which I am while I type this sentence. My favorite place on earth is home with my family. Especially near the end of a 12-hour flight. My 5 top papers: 1“Lewis-Acid Catalysis of a Diels–Alder Reaction in Water”: S. Otto, F. Bertoncin, J. B. F. N. Engberts, J. Am. Chem. Soc. 1996, 118, 7702. (Describes a reaction that I developed at the start of my PhD and that continues to be used in new contexts.) 2“Selection and Amplification of Hosts from Dynamic Combinatorial Libraries of Macrocyclic Disulfides”: S. Otto, R. L. E. Furlan, J. K. M. Sanders, Science 2002, 297, 590. (Dynamic combinatorial chemistry is a powerful tool for developing synthetic receptors.) 3“Systems Chemistry”: R. F. Ludlow, S. Otto, Chem. Soc. Rev. 2008, 37, 101. (Early ideas about the emerging field of systems chemistry.) 4“Mechanosensitive Self-Replication Driven by Self-Organization”: J. M. A. Carnall, C. A. Waudby, A. M. Belenguer, M. C. A. Stuart, J. J.-P. Peyralans, S. Otto, Science 2010, 327, 1502. (A new and in principle general mechanism of self-replication that is driven by self-assembly into nanostructures.) 5“Diversification of self-replicating molecules”: J. W. Sadownik, E. Mattia, P. Nowak, S. Otto, Nature Chem. 2016, 8, 264. (Self-replicating molecules can show behavior akin to the formation of species as encountered in biology.) Volume57, Issue33August 13, 2018Pages 10416-10416 ReferencesRelatedInformation

Existing Self-Replicators Can Direct the Emergence of New Ones.

The study of the interplay between different self-replicating molecules constitutes an important new phase in the synthesis of life and in unravelling the origin of life. Here we show how existing replicators can direct the nature of a newly formed replicator. Starting from the same building block, 6-ring replicators formed when the mixture was exposed to pre-existing 6-membered replicators, while pre-formed 8-membered replicators funneled the building block into 8-ring replicators. Not only ring size, but also the mode of assembly of the rings into stacks was inherited from the pre-existing replicators. These results show that the nature of self-replicating molecules can be strongly influenced by the interplay between different self-replicators, overriding preferences innate to the structure of the building block.

Synthetic Genomics boosts RNA vaccines

Synthetic Genomics has formed two partnerships to pursue RNA replicon-based vaccines, in which self-replicating RNA molecules encode multiple antigen proteins to produce an immune response. One partner, Arcturus Therapeutics, will share its RNA delivery system for an undisclosed payment plus milestones and royalties. Arcturus and Janssen recently agreed to develop hepatitis B treatments with the RNA delivery technology. Synthetic Genomics will also partner with Ceva Sante Animale to develop livestock vaccines.

Self-replicating RNA vaccine functionality modulated by fine-tuning of polyplex delivery vehicle structure

The major limitations with large and complex self-amplifying RNA vaccines (RepRNA) are RNase-sensitivity and inefficient translation in dendritic cells (DCs). Condensing RepRNA with polyethylenimine (PEI) gave positive in vitro readouts, but was largely inferior to virus-like replicon particles (VRP) or direct electroporation. In the present study, we improved such polyplex formulation and determined that fine-tuning of the polyplex structure is essential for ensuring efficacious translation. Thereby, three parameters dominate: (i) PEI molecular weight (MW); (ii) RepRNA:PEI (weight:weight) ratio; and (iii) inclusion of cell penetrating peptides (CPPs). Seven commercially available linear PEIs (MW 2,500–250,000) were classified as strong, intermediate or low for their aptitude at complexing and protecting RepRNA for delivery into porcine blood DCs. Inclusion of (Arg)9 or TAT(57-57) CPPs further modified the translation readouts, but varied for different gene expressions. Dependent on the formulation, translation of the gene of interest (GOI) inserted into the RepRNA (luciferase, or influenza virus hemagglutinin or nucleoprotein) could decrease, while the RepRNA structural gene (E2) translation increased. This was noted in the porcine SK6 cell line, as well as both porcine and, for the first time, human DCs. Two formulations – [Rep/PEI-4,000 (1:3)] and [Rep/PEI-40,000 (1:2)/(Arg)9] were efficacious in vivo in mice and pigs, where specific CD8+ T and CD4+ T-cell responses against the GOI-encoded antigen were observed for the first time. The results demonstrate that different polyplex formulations differ in their interaction with the RepRNA such that only certain genes can be translated. Thus, delivery of these large self-replicating RNA molecules require definition with respect to translation of different genes, rather than just the GOI as is the norm, for identifying optimal delivery for the desired immune activation in vivo.

The origin of a primordial genome through spontaneous symmetry breaking

The heredity of a cell is provided by a small number of non-catalytic templates—the genome. How did genomes originate? Here, we demonstrate the possibility that genome-like molecules arise from symmetry breaking between complementary strands of self-replicating molecules. Our model assumes a population of protocells, each containing a population of self-replicating catalytic molecules. The protocells evolve towards maximising the catalytic activities of the molecules to increase their growth rates. Conversely, the molecules evolve towards minimising their catalytic activities to increase their intracellular relative fitness. These conflicting tendencies induce the symmetry breaking, whereby one strand of the molecules remains catalytic and increases its copy number (enzyme-like molecules), whereas the other becomes non-catalytic and decreases its copy number (genome-like molecules). This asymmetry increases the equilibrium cellular fitness by decreasing mutation pressure and increasing intracellular genetic drift. These results implicate conflicting multilevel evolution as a key cause of the origin of genetic complexity.

Structural and Spectroscopic Properties of Assemblies of Self-Replicating Peptide Macrocycles.

Self-replication at the molecular level is often seen as essential to the early origins of life. Recently a mechanism of self-replication has been discovered in which replicator self-assembly drives the process. We have studied one of the examples of such self-assembling self-replicating molecules to a high level of structural detail using a combination of computational and spectroscopic techniques. Molecular Dynamics simulations of self-assembled stacks of peptide-derived replicators provide insights into the structural characteristics of the system and serve as the basis for semiempirical calculations of the UV–vis, circular dichroism (CD) and infrared (IR) absorption spectra that reflect the chiral organization and peptide secondary structure of the stacks. Two proposed structural models are tested by comparing calculated spectra to experimental data from electron microscopy, CD and IR spectroscopy, resulting in a better insight into the specific supramolecular interactions that lead to self-replication. Specifically, we find a cooperative self-assembly process in which β-sheet formation leads to well-organized structures, while also the aromatic core of the macrocycles plays an important role in the stability of the resulting fibers.

Towards open-ended evolution in self-replicating molecular systems.

In this review we discuss systems of self-replicating molecules in the context of the origin of life and the synthesis of de novo life. One of the important aspects of life is the ability to reproduce and evolve continuously. In this review we consider some of the prerequisites for obtaining unbounded evolution of self-replicating molecules and describe some recent advances in this field. While evolution experiments involving self-replicating molecules have shown promising results, true open-ended evolution has not been realized so far. A full understanding of the requirements for open-ended evolution would provide a better understanding of how life could have emerged from molecular building blocks and what is needed to create a minimal form of life in the laboratory.

Self-Replicating RNA Vaccine Delivery to Dendritic Cells.

Most current vaccines are either inactivated pathogen-derived or protein/peptide-based, although attenuated and vector vaccines have also been developed. The former induce at best moderate protection, even as multimeric antigen, due to limitations in antigen loads and therefore capacity for inducing robust immune defense. While attenuated and vector vaccines offer advantages through their replicative nature, drawbacks and risks remain with potential reversion to virulence and interference from preexisting immunity. New advances averting these problems are combining self-amplifying replicon RNA (RepRNA) technology with nanotechnology. RepRNA are large self-replicating RNA molecules (12-15 kb) derived from viral genomes defective in at least one structural protein gene. They provide sustained antigen production, effectively increasing vaccine antigen payloads over time, without the risk of producing infectious progeny. The major limitation with RepRNA is RNase-sensitivity and inefficient uptake by dendritic cells (DCs)-absolute requirements for efficacious vaccine design. We employed biodegradable delivery vehicles to protect the RepRNA and promote DC delivery. Encapsulating RepRNA into chitosan nanoparticles, as well as condensing RepRNA with polyethylenimine (PEI), cationic lipids, or chitosans, has proven effective for delivery to DCs and induction of immune responses in vivo.

Mechanism of Building Block Exchange in Stacks of Self-Replicating Macrocycles

Self-replicating molecules containing labile reversible covalent bonds are becoming increasingly topical. Yet their robustness to component exchange and the threat of loss of information has received very little attention. We show here that supramolecular interactions can efficiently protect self-replicating molecules from otherwise rapid component exchange.

Diversity of alphasatellites associated with cotton leaf curl disease in Pakistan

BackgroundCotton is a major cash crop of Pakistan and its production is mainly hindered by cotton leaf curl disease (CLCuD). This disease is caused by monopartite begomovirus associated with two satellites named as betasatellite and alphasatellite. Betasatellites are true satellites entirely dependent on helper begomoviruses and are symptom determinants which are essentially required for the typical symptoms of the disease. Alphasatellites are self-replicating circular ssDNA molecules which are associated with CLCuD complex. The role of alphasatellite is not fully understood. ResultCotton samples showing typical CLCuD symptoms were collected from areas across central Punjab, Pakistan during year 2011–12. All samples contained alphasatellites. Mixed-infection of alphasatellites associated with CLCuD complex was documented. Few samples showed the presence of more than one species of alphasatellite. A total of 45 alphasatellites were cloned and sequenced. The size of these alphasatellite ranges from 1362 to 1378bp. All alphasatellites showed three conserved features i.e. 1) A stem-loop structure with a nonanucleotide (TAGTATTAC) sequence (2) An ORF encoding a Rep protein of about 36.6kDa, having up to 315 amino acids (3) An A-rich region of ~200nt. Based on BLAST results we have found six distinct species of alphasatellites namely; Gossypium darwinii symptomless alphasatellite (GDarSLA), Guar leaf curl alphasatellite (GrLCuA), Okra leaf curl alphasatellite (OLCuA), Tomato leaf curl Pakistan alphasatellite (ToLCPKA), Cotton leaf curl Multan alphasatellite (CLCuMA), and Cotton leaf curl Burewala alphasatellite (CLCuBuA). This was also confirmed by phylogenetic analysis. By considering the species cut-off limit for alphasatellites (83%) the isolates fall into 5 species. But the percentage identity between some CLCuBuA and CLCuMA was 83.3, so they are proposed to be considered as two different species. ConclusionOur data shows that at least six species of alphasatellites are found associated with cotton leaf curl disease in Pakistan. Field samples are often associated with multiple species and one sample was found associated with three distinct alphasatellites in a single plant under field conditions. Infection of multiple alphasatellite and their probable role in CLCuD are discussed.