Nature 459, 239-242 (14 May 2009) | doi:10.1038/nature08013; Received 11 December 2008; Accepted 24 March 2009
Synthesis of activated pyrimidine ribonucleotides in prebiotically plausible conditions
Matthew W. Powner1, Béatrice Gerland1 & John D. Sutherland1
1. School of Chemistry, The University of Manchester, Oxford Road, Manchester M13 9PL, UK
Correspondence to: John D. Sutherland1 Correspondence and requests for materials should be addressed to J.D.S. (Email: john.sutherland@manchester.ac.uk).
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Abstract
At some stage in the origin of life, an informational polymer must have arisen by purely chemical means. According to one version of the 'RNA world' hypothesis1, 2, 3 this polymer was RNA, but attempts to provide experimental support for this have failed4, 5. In particular, although there has been some success demonstrating that 'activated' ribonucleotides can polymerize to form RNA6, 7, it is far from obvious how such ribonucleotides could have formed from their constituent parts (ribose and nucleobases). Ribose is difficult to form selectively8, 9, and the addition of nucleobases to ribose is inefficient in the case of purines10 and does not occur at all in the case of the canonical pyrimidines11. Here we show that activated pyrimidine ribonucleotides can be formed in a short sequence that bypasses free ribose and the nucleobases, and instead proceeds through arabinose amino-oxazoline and anhydronucleoside intermediates. The starting materials for the synthesis—cyanamide, cyanoacetylene, glycolaldehyde, glyceraldehyde and inorganic phosphate—are plausible prebiotic feedstock molecules12, 13, 14, 15, and the conditions of the synthesis are consistent with potential early-Earth geochemical models. Although inorganic phosphate is only incorporated into the nucleotides at a late stage of the sequence, its presence from the start is essential as it controls three reactions in the earlier stages by acting as a general acid/base catalyst, a nucleophilic catalyst, a pH buffer and a chemical buffer. For prebiotic reaction sequences, our results highlight the importance of working with mixed chemical systems in which reactants for a particular reaction step can also control other steps.

Life’s First Spark Re-Created in the Laboratory

A fundamental but elusive step in the early evolution of life on Earth has been replicated in a laboratory.
Researchers synthesized the basic ingredients of RNA, a molecule from which the simplest self-replicating structures are made. Until now, they couldn’t explain how these ingredients might have formed.
“It’s like molecular choreography, where the molecules choreograph their own behavior,” said organic chemist John Sutherland of the University of Manchester, co-author of a study in Nature Wednesday.
RNA is now found in living cells, where it carries information between genes and protein-manufacturing cellular components. Scientists think RNA existed early in Earth’s history, providing a necessary intermediate platform between pre-biotic chemicals and DNA, its double-stranded, more-stable descendant.

However, though researchers have been able to show how RNA’s component molecules, called ribonucleotides, could assemble into RNA, their many attempts to synthesize these ribonucleotides have failed. No matter how they combined the ingredients — a sugar, a phosphate, and one of four different nitrogenous molecules, or nucleobases — ribonucleotides just wouldn’t form.

Sutherland’s team took a different approach in what Harvard molecular biologist Jack Szostak called a “synthetic tour de force” in an accompanying commentary in Nature.

“By changing the way we mix the ingredients together, we managed to make ribonucleotides,” said Sutherland. “The chemistry works very effectively from simple precursors, and the conditions required are not distinct from what one might imagine took place on the early Earth.”

Like other would-be nucleotide synthesizers, Sutherland’s team included phosphate in their mix, but rather than adding it to sugars and nucleobases, they started with an array of even simpler molecules that were probably also in Earth’s primordial ooze.

They mixed the molecules in water, heated the solution, then allowed it to evaporate, leaving behind a residue of hybrid, half-sugar, half-nucleobase molecules. To this residue they again added water, heated it, allowed it evaporate, and then irradiated it.

At each stage of the cycle, the resulting molecules were more complex. At the final stage, Sutherland’s team added phosphate. “Remarkably, it transformed into the ribonucleotide!” said Sutherland.

According to Sutherland, these laboratory conditions resembled those of the life-originating “warm little pond” hypothesized by Charles Darwin if the pond “evaporated, got heated, and then it rained and the sun shone.”
Such conditions are plausible, and Szostak imagined the ongoing cycle of evaporation, heating and condensation providing “a kind of organic snow which could accumulate as a reservoir of material ready for the next step in RNA synthesis.”

Intriguingly, the precursor molecules used by Sutherland’s team have been identified in interstellar dust clouds and on meteorites.

“Ribonucleotides are simply an expression of the fundamental principles of organic chemistry,” said Sutherland. “They’re doing it unwittingly. The instructions for them to do it are inherent in the structure of the precursor materials. And if they can self-assemble so easily, perhaps they shouldn’t be viewed as complicated.”

Michael C wrote:Abiogenesis (life forming from non-living material)

Tilokarat wrote:A biology teacher at the high school where I work during the day said that someone will raise the question of whether any of the test materials were contaminated with bacteria, which would provide 'life' in the experiment results.

westerby wrote:

Michael C wrote:Abiogenesis (life forming from non-living material)

Bringing it down to Wester's level, it means that we are one step closer to author, Philip K Dick's concept of the Replicant - ever seen Blade Runner?

Chemist Shows How RNA Can Be the Starting Point for Life

An English chemist has found the hidden gateway to the RNA world, the chemical milieu from which the first forms of life are thought to have emerged on earth some 3.8 billion years ago.

He has solved a problem that for 20 years has thwarted researchers trying to understand the origin of life — how the building blocks of RNA, called nucleotides, could have spontaneously assembled themselves in the conditions of the primitive earth. The discovery, if correct, should set researchers on the right track to solving many other mysteries about the origin of life. It will also mean that for the first time a plausible explanation exists for how an information-carrying biological molecule could have emerged through natural processes from chemicals on the primitive earth.

The author, John D. Sutherland, a chemist at the University of Manchester, likened his work to a crossword puzzle in which doing the first clues makes the others easier. “Whether we’ve done one across is an open question,” he said. “Our worry is that it may not be right.”

Other researchers say they believe he has made a major advance in prebiotic chemistry, the study of the natural chemical reactions that preceded the first living cells. “It is precisely because this work opens up so many new directions for research that it will stand for years as one of the great advances in prebiotic chemistry,” Jack Szostak of the Massachusetts General Hospital wrote in a commentary in Nature, where the work is being published on Thursday.

Scientists have long suspected that the first forms of life carried their biological information not in DNA but in RNA, its close chemical cousin. Though DNA is better known because of its storage of genetic information, RNA performs many of the trickiest operations in living cells. RNA seems to have delegated the chore of data storage to the chemically more stable DNA eons ago. If the first forms of life were based on RNA, then the issue is to explain how the first RNA molecules were formed.

For more than 20 years researchers have been working on this problem. The building blocks of RNA, known as nucleotides, each consist of a chemical base, a sugar molecule called ribose and a phosphate group. Chemists quickly found plausible natural ways for each of these constituents to form from natural chemicals. But there was no natural way for them all to join together.

The spontaneous appearance of such nucleotides on the primitive earth “would have been a near miracle,” two leading researchers, Gerald Joyce and Leslie Orgel, wrote in 1999. Others were so despairing that they believed some other molecule must have preceded RNA and started looking for a pre-RNA world.

The miracle seems now to have been explained. In the article in Nature, Dr. Sutherland and his colleagues Matthew W. Powner and Béatrice Gerland report that they have taken the same starting chemicals used by others but have caused them to react in a different order and in different combinations than in previous experiments. they discovered their recipe, which is far from intuitive, after 10 years of working through every possible combination of starting chemicals.

Instead of making the starting chemicals form a sugar and a base, they mixed them in a different order, in which the chemicals naturally formed a compound that is half-sugar and half-base. When another half-sugar and half-base are added, the RNA nucleotide called ribocytidine phosphate emerges.

A second nucleotide is created if ultraviolet light is shined on the mixture. Dr. Sutherland said he had not yet found natural ways to generate the other two types of nucleotides found in RNA molecules, but synthesis of the first two was thought to be harder to achieve.

If all four nucleotides formed naturally, they would zip together easily to form an RNA molecule with a backbone of alternating sugar and phosphate groups. The bases attached to the sugar constitute a four-letter alphabet in which biological information can be represented.

“My assumption is that we are here on this planet as a fundamental consequence of organic chemistry,” Dr. Sutherland said. “So it must be chemistry that wants to work.”

The reactions he has described look convincing to most other chemists. “The chemistry is very robust — all the yields are good and the chemistry is simple,” said Dr. Joyce, an expert on the chemical origin of life at the Scripps Research Institute in La Jolla, Calif.

Self-Sustained Replication of an RNA Enzyme
Tracey A. Lincoln and Gerald F. Joyce*

An RNA enzyme that catalyzes the RNA-templated joining of RNA was converted to a format whereby two enzymes catalyze each other's synthesis from a total of four oligonucleotide substrates. These cross-replicating RNA enzymes undergo self-sustained exponential amplification in the absence of proteins or other biological materials. Amplification occurs with a doubling time of about 1 hour and can be continued indefinitely. Populations of various cross-replicating enzymes were constructed and allowed to compete for a common pool of substrates, during which recombinant replicators arose and grew to dominate the population. These replicating RNA enzymes can serve as an experimental model of a genetic system. Many such model systems could be constructed, allowing different selective outcomes to be related to the underlying properties of the genetic system.

Department of Chemistry, Department of Molecular Biology, and the Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.

Source:
Science 27 February 2009:
Vol. 323. no. 5918, pp. 1229 - 1232

How Did Life Begin? RNA That Replicates Itself Indefinitely Developed For First Time

One of the most enduring questions is how life could have begun on Earth. Molecules that can make copies of themselves are thought to be crucial to understanding this process as they provide the basis for heritability, a critical characteristic of living systems. New findings could inform biochemical questions about how life began.

Now, a pair of Scripps Research Institute scientists has taken a significant step toward answering that question. The scientists have synthesized for the first time RNA enzymes that can replicate themselves without the help of any proteins or other cellular components, and the process proceeds indefinitely.

The work was recently published in the journal Science.

In the modern world, DNA carries the genetic sequence for advanced organisms, while RNA is dependent on DNA for performing its roles such as building proteins. But one prominent theory about the origins of life, called the RNA World model, postulates that because RNA can function as both a gene and an enzyme, RNA might have come before DNA and protein and acted as the ancestral molecule of life. However, the process of copying a genetic molecule, which is considered a basic qualification for life, appears to be exceedingly complex, involving many proteins and other cellular components.

For years, researchers have wondered whether there might be some simpler way to copy RNA, brought about by the RNA itself. Some tentative steps along this road had previously been taken by the Joyce lab and others, but no one could demonstrate that RNA replication could be self-propagating, that is, result in new copies of RNA that also could copy themselves.

In Vitro Evolution

A few years after Tracey Lincoln arrived at Scripps Research from Jamaica to pursue her Ph.D., she began exploring the RNA-only replication concept along with her advisor, Professor Gerald Joyce, M.D., Ph.D., who is also Dean of the Faculty at Scripps Research. Their work began with a method of forced adaptation known as in vitro evolution. The goal was to take one of the RNA enzymes already developed in the lab that could perform the basic chemistry of replication, and improve it to the point that it could drive efficient, perpetual self-replication.

Lincoln synthesized in the laboratory a large population of variants of the RNA enzyme that would be challenged to do the job, and carried out a test-tube evolution procedure to obtain those variants that were most adept at joining together pieces of RNA.

Ultimately, this process enabled the team to isolate an evolved version of the original enzyme that is a very efficient replicator, something that many research groups, including Joyce's, had struggled for years to obtain. The improved enzyme fulfilled the primary goal of being able to undergo perpetual replication. "It kind of blew me away," says Lincoln.

Immortalizing Molecular Information

The replicating system actually involves two enzymes, each composed of two subunits and each functioning as a catalyst that assembles the other. The replication process is cyclic, in that the first enzyme binds the two subunits that comprise the second enzyme and joins them to make a new copy of the second enzyme; while the second enzyme similarly binds and joins the two subunits that comprise the first enzyme. In this way the two enzymes assemble each other — what is termed cross-replication. To make the process proceed indefinitely requires only a small starting amount of the two enzymes and a steady supply of the subunits.

"This is the only case outside biology where molecular information has been immortalized," says Joyce.

Not content to stop there, the researchers generated a variety of enzyme pairs with similar capabilities. They mixed 12 different cross-replicating pairs, together with all of their constituent subunits, and allowed them to compete in a molecular test of survival of the fittest. Most of the time the replicating enzymes would breed true, but on occasion an enzyme would make a mistake by binding one of the subunits from one of the other replicating enzymes. When such "mutations" occurred, the resulting recombinant enzymes also were capable of sustained replication, with the most fit replicators growing in number to dominate the mixture. "To me that's actually the biggest result," says Joyce.

The research shows that the system can sustain molecular information, a form of heritability, and give rise to variations of itself in a way akin to Darwinian evolution. So, says Lincoln, "What we have is non-living, but we've been able to show that it has some life-like properties, and that was extremely interesting."

Knocking on the Door of Life

The group is pursuing potential applications of their discovery in the field of molecular diagnostics, but that work is tied to a research paper currently in review, so the researchers can't yet discuss it.

But the main value of the work, according to Joyce, is at the basic research level. "What we've found could be relevant to how life begins, at that key moment when Darwinian evolution starts." He is quick to point out that, while the self-replicating RNA enzyme systems share certain characteristics of life, they are not themselves a form of life.

The historical origin of life can never be recreated precisely, so without a reliable time machine, one must instead address the related question of whether life could ever be created in a laboratory. This could, of course, shed light on what the beginning of life might have looked like, at least in outline. "We're not trying to play back the tape," says Lincoln of their work, "but it might tell us how you go about starting the process of understanding the emergence of life in the lab."

Joyce says that only when a system is developed in the lab that has the capability of evolving novel functions on its own can it be properly called life. "We're knocking on that door," he says, "But of course we haven't achieved that."

The subunits in the enzymes the team constructed each contain many nucleotides, so they are relatively complex and not something that would have been found floating in the primordial ooze. But, while the building blocks likely would have been simpler, the work does finally show that a simpler form of RNA-based life is at least possible, which should drive further research to explore the RNA World theory of life's origins.

Journal reference: 1. Lincoln et al. Self-Sustained Replication of an RNA Enzyme. Science, Jan 8, 2009; DOI: 10.1126/science.1167856

Source: Science Daily