A Magnesium Binding Site And The Anomeric Effect Regulate The Abiotic Redox Chemistry Of Nicotinamide Nucleotides.

Nicotinamide adenine dinucleotide (NAD+) is a redox active molecule that is universally found in biology. Despite the importance and simplicity of this molecule, few reports exist that investigate which molecular features are important for the activity of this ribodinucleotide. By exploiting the nonenzymatic reduction and oxidation of NAD+ by pyruvate and methylene blue, respectively, we were able to identify key molecular features necessary for the intrinsic activity of NAD+ through kinetic analysis. Such features may explain how NAD+ could have been selected early during the emergence of life. Simpler molecules, such as nicotinamide, that lack an anomeric carbon are incapable of accepting electrons from pyruvate. The phosphate moiety inhibits activity in the absence of metal ions but facilitates activity at physiological pH and model prebiotic conditions by recruiting catalytic Mg2+. Reduction proceeds through consecutive single electron transfer events. Of the derivatives tested, including nicotinamide mononucleotide, nicotinamide riboside, 3-(aminocarbonyl)-1-(2,3-dihydroxypropyl)pyridinium, 1-methylnicotinamide, and nicotinamide, only NAD+ and nicotinamide mononucleotide would be capable of efficiently accepting and donating electrons within a nonenzymatic electron transport chain. The data are consistent with early metabolic chemistry exploiting NAD+ or nicotinamide mononucleotide and not simpler molecules.

Prebiotic thiol-catalyzed thioamide bond formation.

Thioamide bonds are important intermediates in prebiotic chemistry. In cyanosulfidic prebiotic chemistry, they serve as crucial intermediates in the pathways that lead to the formation of many important biomolecules (e.g., amino acids). They can also serve as purine and pyrimidine precursors, the two classes of heterocycle employed in genetic molecules. Despite their importance, the formation of thioamide bonds from nitriles under prebiotic conditions has required large excesses of sulfide or compounds with unknown prebiotic sources. Here, we describe the thiol-catalyzed formation of thioamide bonds from nitriles. We show that the formation of the simplest of these compounds, thioformamide, forms readily in spark-discharge experiments from hydrogen cyanide, sulfide, and a methanethiol catalyst, suggesting potential accumulation on early Earth. Lastly, we demonstrate that thioformamide has a Gibbs energy of hydrolysis ( Δ G r ∘ ) comparable to other energy-currencies on early Earth such as pyrophosphate and thioester bonds. Overall, our findings imply that thioamides might have been abundant on early Earth and served a variety of functions during chemical evolution.

Ring-Closure on the Rocks in a Prebiotic Environment.

Ring-closure is a key step in current pyrimidine anabolism and one may wonder whether cyclisation reactions could be promoted in the geochemical context at the origins of life, i. e. with the help of minerals. Various prebiotic minerals were tested in this work, including silica, carbonates, microporous minerals. In particular, the role of zinc ions supported on minerals was investigated in view of its presence in the catalytic site of cyclic amidohydrolase enzymes. Based on in situ (TGA: ThermoGravimetric Analysis, ATR-IR: Attenuated Total Reflectance-InfraRed) and ex situ (1 H NMR- Nuclear Magnetic Resonance) characterisations, we identified the products of thermal activation of NCA (N-carbamoyl-aspartic acid) in wetting-and-drying scenarios on the surface of minerals. NCA can cyclize extensively only on some surfaces, with the predominant product being 5-carboxymethylhydantoin (Hy) rather than dihydroorotate (DHO), while there is a competition with hydrolysis on others. Replacing the enzymes with heterogeneous catalysts also works with other reactions catalysed by enzymes of the cyclic amidohydrolases family. The role of the hydrophilicity/hydrophobicity of minerals as well as the regioselectivity of the cyclisation (5-carboxymethylhydantoin versus dihydroorotate) are examined.

The limits of metabolic heredity in protocells.

The universal core of metabolism could have emerged from thermodynamically favoured prebiotic pathways at the origin of life. Starting with H2 and CO2, the synthesis of amino acids and mixed fatty acids, which self-assemble into protocells, is favoured under warm anoxic conditions. Here, we address whether it is possible for protocells to evolve greater metabolic complexity, through positive feedbacks involving nucleotide catalysis. Using mathematical simulations to model metabolic heredity in protocells, based on branch points in protometabolic flux, we show that nucleotide catalysis can indeed promote protocell growth. This outcome only occurs when nucleotides directly catalyse CO2 fixation. Strong nucleotide catalysis of other pathways (e.g. fatty acids and amino acids) generally unbalances metabolism and slows down protocell growth, and when there is competition between catalytic functions cell growth collapses. Autocatalysis of nucleotide synthesis can promote growth but only if nucleotides also catalyse CO2 fixation; autocatalysis alone leads to the accumulation of nucleotides at the expense of CO2 fixation and protocell growth rate. Our findings offer a new framework for the emergence of greater metabolic complexity, in which nucleotides catalyse broad-spectrum processes such as CO2 fixation, hydrogenation and phosphorylation important to the emergence of genetic heredity at the origin of life.

Protometabolism as out-of-equilibrium chemistry.

It is common to compare life with machines. Both consume fuel and release waste to run. In biology, the engine that drives the living system is referred to as metabolism. However, attempts at deciphering the origins of metabolism do not focus on this energetic relationship that sustains life but rather concentrate on nonenzymatic reactions that produce all the intermediates of an extant metabolic pathway. Such an approach is akin to studying the molecules produced from the burning of coal instead of deciphering how the released energy drives the movement of pistons and ultimately the train when investigating the mechanisms behind locomotion. Theories that do explicitly invoke geological chemical gradients to drive metabolism most frequently feature hydrothermal vent conditions, but hydrothermal vents are not the only regions of the early Earth that could have provided the fuel necessary to sustain the Earth's first (proto)cells. Here, we give examples of prior reports on protometabolism and highlight how more recent investigations of out-of-equilibrium systems may point to alternative scenarios more consistent with the majority of prebiotic chemistry data accumulated thus far. This article is part of the theme issue 'Emergent phenomena in complex physical and socio-technical systems: from cells to societies'.

Emergence of a Promiscuous Peroxidase Under Non-Equilibrium Conditions.

Herein, we report the substrate induced generation of a transient catalytic microenvironment from a single amino acid functionalized fatty acid in presence of a cofactor hemin. The catalytic state accessed under non-equilibrium conditions showed acceleration of peroxidase activity resulting in degradation of the substrate and subsequently led to disassembly. Equilibrated systems could not access the three-dimensional microphases and showed substantially lower catalytic activity. Further, the assembled state showed latent catalytic function (promiscuity) to hydrolyze a precursor to yield the same substrate. Consequently, the assembly demonstrated protometabolism by exploiting the peroxidase-hydrolase cascade to augment the lifetime and the mechanical properties of the catalytic state.

Coenzymes and Their Role in the Evolution of Life.

The evolution of coenzymes, or their impact on the origin of life, is fundamental for understanding our own existence. Having established reasonable hypotheses about the emergence of prebiotic chemical building blocks, which were probably created under palaeogeochemical conditions, and surmising that these smaller compounds must have become integrated to afford complex macromolecules such as RNA, the question of coenzyme origin and its relation to the evolution of functional biochemistry should gain new impetus. Many coenzymes have a simple chemical structure and are often nucleotide-derived, which suggests that they may have coexisted with the emergence of RNA and may have played a pivotal role in early metabolism. Based on current theories of prebiotic evolution, which attempt to explain the emergence of privileged organic building blocks, this Review discusses plausible hypotheses on the prebiotic formation of key elements within selected extant coenzymes. In combination with prebiotic RNA, coenzymes may have dramatically broadened early protometabolic networks and the catalytic scope of RNA during the evolution of life.

The optimal size of protocells from simple entropic considerations.

Potential constraints on protocell size are developed from simple entropic considerations. To do that, two new different indexes as measures of their structural and dynamic order were developed and applied to an elemental model of the heterotrophic protocell. According to our results, cell size should be a key factor determining the potential of these primitive systems to evolve and consequently to support life. Our analyses also suggest that the size of the optimal vesicles could be constrained to have radii in the interval [Formula: see text], where the two extreme limits [Formula: see text] and [Formula: see text] represent the states of maximum structural order (largest accumulation of substrate inside the vesicle) and the maximum flux of entropy production, respectively. According to the above criteria, the size of the optimum vesicles falls, approximately, in the same spatial range estimated for biological living cells assuming plausible values for the second-order rate constant involved in the catabolic process. Furthermore, the existence of very small vesicles could be seriously affected by the limited efficiency, far from the theoretical limits, with which these catabolic processes may proceed in a prebiotic system.

The Role of Aqueous Aerosols in the "Glyoxylate Scenario": An Experimental Approach.

The origin of life is one of the fundamental questions in science. Eschenmoser proposed the "glyoxylate scenario", in which plausible abiotic synthesis pathways were suggested to be compatible with the constraints of prebiotic chemistry. In this proposal, the stem compound is HCN. In this work, we explore the "glyoxylate scenario" through several syntheses of HCN polymers, paying particular attention to the role of the aqueous aerosols, together with statistical methods, as a step to elucidate the synthetic problem of the origin of life. The soluble and insoluble HCN polymers synthetized were analyzed by GC-MS. We identified, for the first time, glyoxylic acid in these polymers, together with some constituents of the reductive tricarboxylic acid cycle, amino acids and several N-heterocycles. The findings presented herein, as the first global approach to the "glyoxylate scenario", give full effect to this hypothesis and prove that aqueous aerosols could play an important role in this plausible scene of the origin of life.

Prebiotic Synthesis of Aspartate Using Life's Metabolism as a Guide.

A protometabolic approach to the origins of life assumes that the conserved biochemistry of metabolism has direct continuity with prebiotic chemistry. One of the most important amino acids in modern biology is aspartic acid, serving as a nodal metabolite for the synthesis of many other essential biomolecules. Aspartate's prebiotic synthesis is complicated by the instability of its precursor, oxaloacetate. In this paper, we show that the use of the biologically relevant cofactor pyridoxamine, supported by metal ion catalysis, is sufficiently fast to offset oxaloacetate's degradation. Cu2+-catalysed transamination of oxaloacetate by pyridoxamine achieves around a 5% yield within 1 h, and can operate across a broad range of pH, temperature, and pressure. In addition, the synthesis of the downstream product ß-alanine may also take place in the same reaction system at very low yields, directly mimicking an archaeal synthesis route. Amino group transfer supported by pyridoxal is shown to take place from aspartate to alanine, but the reverse reaction (alanine to aspartate) shows a poor yield. Overall, our results show that the nodal metabolite aspartate and related amino acids can indeed be synthesised via protometabolic pathways that foreshadow modern metabolism in the presence of the simple cofactor pyridoxamine and metal ions.

Do Soluble Phosphates Direct the Formose Reaction towards Pentose Sugars?

The formose reaction has been a leading hypothesis for the prebiotic synthesis of sugars such as ribose for many decades but tends to produce complex mixtures of sugars and often tars. Channeling the formose reaction towards the synthesis of biologically useful sugars such as ribose has been a holy grail of origins-of-life research. Here, we tested the hypothesis that a simple, prebiotically plausible phosphorylating agent, acetyl phosphate, could direct the formose reaction towards ribose through phosphorylation of intermediates in a manner resembling gluconeogenesis and the pentose phosphate pathway. We did indeed find that addition of acetyl phosphate to a developing formose reaction stabilized pentoses, including ribose, such that after 5 h of reaction about 10-fold more ribose remained compared with control runs. But mechanistic analyses using liquid chromatography-mass spectrometry showed that, far from being directed towards ribose by phosphorylation, the formose reaction was halted by the precipitation of Ca2+ ions as phosphate minerals such as apatite and hydroxyapatite. Adding orthophosphate had the same effect. Phosphorylated sugars were only detected below the limit of quantification when adding acetyl phosphate. Nonetheless, our findings are not strictly negative. The sensitivity of the formose reaction to geochemically reasonable conditions, combined with the apparent stability of ribose under these conditions, serves as a valuable constraint on possible pathways of sugar synthesis at the origin of life.

Thermodynamics of Potential CHO Metabolites in a Reducing Environment.

How did metabolism arise and evolve? What chemical compounds might be suitable to support and sustain a proto-metabolism before the advent of more complex co-factors? We explore these questions by using first-principles quantum chemistry to calculate the free energies of CHO compounds in aqueous solution, allowing us to probe the thermodynamics of core extant cycles and their closely related chemical cousins. By framing our analysis in terms of the simplest feasible cycle and its permutations, we analyze potentially favorable thermodynamic cycles for CO2 fixation with H2 as a reductant. We find that paying attention to redox states illuminates which reactions are endergonic or exergonic. Our results highlight the role of acetate in proto-metabolic cycles, and its connection to other prebiotic molecules such as glyoxalate, glycolaldehyde, and glycolic acid.

Prebiotic Reaction Networks in Water.

A prevailing strategy in origins of life studies is to explore how chemistry constrained by hypothetical prebiotic conditions could have led to molecules and system level processes proposed to be important for life's beginnings. This strategy has yielded model prebiotic reaction networks that elucidate pathways by which relevant compounds can be generated, in some cases, autocatalytically. These prebiotic reaction networks provide a rich platform for further understanding and development of emergent "life-like" behaviours. In this review, recent advances in experimental and analytical procedures associated with classical prebiotic reaction networks, like formose and Miller-Urey, as well as more recent ones are highlighted. Instead of polymeric networks, i.e., those based on nucleic acids or peptides, the focus is on small molecules. The future of prebiotic chemistry lies in better understanding the genuine complexity that can result from reaction networks and the construction of a centralised database of reactions useful for predicting potential network evolution is emphasised.

Phosphorylation and acylation transfer reactions: Clues to a dual origin of metabolism.

Many theories of the origin of life focus on only one primitive polymer as an archetype of a world paradigm. However, life would have emerged within more complex scenarios where a variety of molecules and diverse polymers interconnected by a few similar chemical reactions. Previous work suggested that the ancestors of all major biopolymers would have arisen from abiotic template independent replication processes. They would have been organized in two closed sets of polymerization cycles: polysaccharides, polyribonucleotides and polyphosphates on one site; and peptides, fatty acids and polyhydroxyalkanoates on the other site. Then, these polymerization reaction cycles integrated into a minimal organization closure. Here, the purpose was to explore which kind of reactions could have supported the chemical networks that led to the early (bio)polymers. As a result, the proposed overview suggests that phosphorylation and acylation transfer reactions would have arisen independently and forged two distinct chemical systems that provided the phosphorylated and carboxylated intermediates used for the synthesis of the corresponding polymers. In this sense, modern metabolism may still reflect its dual nature, probably relying on these two reaction networks from the beginnings.

Natural Radioactive Environments as Sources of Local Disequilibrium for the Emergence of Life.

Certain subterranean environments of Earth have naturally accumulated long-lived radionuclides, such as 238U, 232Th, and 40K, near the presence of liquid water. In these natural radioactive environments, water radiolysis can produce chemical species of biological importance, such as H2. Although the proposal of radioactive decay as an alternative source of energy for living systems has existed for >30 years, this hypothesis gained strength after the recent discovery of a peculiar ecosystem in a gold mine in South Africa, whose existence is dependent on chemical species produced by water radiolysis. In this study, we calculate the chemical disequilibrium generated locally by water radiolysis due to gamma radiation. We then analyze the possible contribution of this disequilibrium for the emergence of life, considering conditions of early Earth and having as reference the alkaline hydrothermal vent theory. Results from our kinetic model point out the similarities between the conditions caused by water radiolysis and those found on alkaline hydrothermal systems. Our model produces a steady increase of pH with time, which favors the formation of a natural electrochemical gradient and the precipitation of minerals with catalytic activity for protometabolism in this aqueous environment. We conclude by describing a possible free-energy conversion mechanism based on protometabolism, which could be a requisite for the emergence of life in Hadean Earth.

The ambivalent role of water at the origins of life.

Life as we know it would not exist without water. However, water molecules not only serve as a solvent and reactant but can also promote hydrolysis, which counteracts the formation of essential organic molecules. This conundrum constitutes one of the central issues in origin of life. Hydrolysis is an important part of energy metabolism for all living organisms but only because, inside cells, it is a controlled reaction. How could hydrolysis have been regulated under prebiotic settings? Lower water activities possibly provide an answer: geochemical sites with less free and more bound water can supply the necessary conditions for protometabolic reactions. Such conditions occur in serpentinising systems, hydrothermal sites that synthesise hydrogen gas via rock-water interactions. Here, we summarise the parallels between biotic and abiotic means of controlling hydrolysis in order to narrow the gap between biochemical and geochemical reactions and briefly outline how hydrolysis could even have played a constructive role at the origin of molecular self-organisation.

The Messy Alkaline Formose Reaction and Its Link to Metabolism.

Sugars are essential for the formation of genetic elements such as RNA and as an energy/food source. Thus, the formose reaction, which autocatalytically generates a multitude of sugars from formaldehyde, has been viewed as a potentially important prebiotic source of biomolecules at the origins of life. When analyzing our formose solutions we find that many of the chemical species are simple carboxylic acids, including α-hydroxy acids, associated with metabolism. In this work we posit that the study of the formose reaction, under alkaline conditions and moderate hydrothermal temperatures, should not be solely focused on sugars for genetic materials, but should focus on the origins of metabolism (via metabolic molecules) as well.

Progress in synthesizing protocells.

IMPACT STATEMENT: Advances in the understanding of the biophysics of membranes, the nonenzymatic and enzymatic polymerization of RNA, and in the design of complex chemical reaction networks have led to a new, integrated way of viewing the shared chemistry needed to sustain life. Although a protocell capable of Darwinian evolution has yet to be built, the seemingly disparate pieces are beginning to fit together. At the very least, better cellular mimics are on the horizon that will likely teach us much about the physicochemical underpinnings of cellular life.