RNA as the Central Molecule for a Prebiotic Metabolism

Extant cellular life is based on well understood principles of molecular biology that comprise the protein-dependent replication of nucleic acids and the nucleic acid-dependent encoding of proteins. Although it is obvious that a simpler system must have preceded today's complex cellular biology, this interdependence of proteins and nucleic acids has long represented a profound puzzle (often depicted as the "chicken and egg" dilemma) for researchers attempting to determine the nature of a primitive "biochemical" metabolism. In fact, a "satisfactory" answer to this dilemma could not be envisioned until the RNA-World concept was first hypothesized in the1960s by F.H.C. Crick6 and L.E. Orgel7 among others and later named by W. Gilbert.8 These authors postulated an autonomous RNA-based "organism" or protocell in which RNA strands could perform several functions presently carried out by proteins, first and foremost that of RNA replication, while acting as genetic information repository. At that time, no catalytic RNAs (so called ribozymes) had been discovered and this idea was rather speculative because it was only supported by the ubiquitous distribution of coenzymes incorporating nucleotides in their structures. Natural selection through replication and mutation was also known to be the sole mechanism for the evolution of complex biochemical systems from simpler ones. The discovery of self-splicing ribozymes in contemporary cells and more recently the demonstration that ubiquitous, ribosomal peptide synthesis is a ribozyme-catalyzed reaction9 have lent increased plausibility to the idea of an RNA World as the precursor of the current DNA-RNA-Protein World. Although the RNA World seems inevitable at some point during the emergence ofcellular Life from inanimate matter, it is not clear whether it represented the first system during evolution towards Life. In fact, the evidence at hand still leaves open questions about the origins and "biochemistry" of the RNA World.

The realization of a functional RNA World requires a series of events (Fig. 10.1) including the abiotic synthesis ofRNA monomers and their assembly into oligomers (in the likely presence of metal catalysts) that could be elongated either by monomers or by ligation with another oligomer. These molecules would have had to serve as templates for their own copying or replication and all these events must have initially occurred non-enzymatically. At that stage, a pool of RNAs (a large set of RNA molecules) must have existed from which a set of catalytic RNAs (among them, RNA-based RNA copying molecules) emerged through natural selection that together sustained exponential growth in the prebiotic environment.

(i) Implications of the RNA Activity for RNA Polymerization

Catalysis requires molecular recognition of a target molecule by the catalyst. It is likely that early ribozymes were not as substrate specific as today's protein enzymes, a fact that would have allowed them to take full advantage ofthe various substrate derivatives synthesized abiotically while still permitting a relatively efficient catalysis. Just as for protein enzymes, the recognition oftarget molecules by ribozymes is determined by the conformation ofthe RNA molecules (i.e., their three-dimensional shape) (Fig. 10.2 and Box 10.1).

The assembly of RNA motifs requires relatively long polymers: known synthetic or natural RNA polymers capable of catalysis or molecular recognition tend to be composed of at least 20-30 units with a large nucleobase fraction involved in the formation of structural motifs. In vitro selected RNA ligases and polymerases are formed by even longer strands of at least 50 and 150 monomers, respectively.

Such lengths have implications for the RNA-polymerization process. Indeed, ifa 50-mer RNA is needed for a given activity, 450 or 1030 different RNA sequences (assuming four different nucleobases) theoretically exist, but 3.5 x 107 kg RNA (the total weight of all possible molecules of that length) cannot likely be made at once. Although a significant number of such sequences could have had the same activity, thereby reducing the above amount, concentration issues must not be overlooked: a threshold concentration of active molecules is always necessary for a given activity and must be taken into account when considering an initiation of the RNA World. This is especially true if several sequences were needed simultaneously in a cooperative role. Thus, the RNA polymerization

*Pierre-Alain Monnard—Los Alamos National Laboratory, Earth and Environmental Science Division (EES-6), Los Alamos, USA. Current address: Flint Center, University of Southern Denmark, Odense M, Denmark. Email: [email protected]

Figure 10.1. Hypothetical route to the de novo RNA World. N stands for any nucleobase, (pN) for a RNA oligomer or polymer. m, m+1 and m+n subscripts refer to the length of a polymer. A) Synthesis of the RNA monomers from simple organic precursors. This phase relies on prebiotic chemistry. B) Oligomerization of the monomers. Monomers are incorporated (polymerized) into oligomers in the likely presence of metal-ion catalysts as no protein enzymes were present to catalyze such a reaction. C) Synthesis of longer polymers. Polymers required for catalytic activity can be obtained either by ligation of previously formed oligomers or further monomer addition. D) Non-enzymatic, template-directed polymerization. To transmit the sequence of catalytic RNA fragments, the ability to non-enzymatically copy these molecules is essential. This process will not only permit an increase of the catalytic activity, but also of the RNA-pool size as well as the emergence of new catalysts because of copying errors. E) The RNA World. When the RNA pool contains molecules that are also catalysts of their own copying, as well as those of any other functional RNAs, a full-fledged RNA World is achieved.

Figure 10.1. Hypothetical route to the de novo RNA World. N stands for any nucleobase, (pN) for a RNA oligomer or polymer. m, m+1 and m+n subscripts refer to the length of a polymer. A) Synthesis of the RNA monomers from simple organic precursors. This phase relies on prebiotic chemistry. B) Oligomerization of the monomers. Monomers are incorporated (polymerized) into oligomers in the likely presence of metal-ion catalysts as no protein enzymes were present to catalyze such a reaction. C) Synthesis of longer polymers. Polymers required for catalytic activity can be obtained either by ligation of previously formed oligomers or further monomer addition. D) Non-enzymatic, template-directed polymerization. To transmit the sequence of catalytic RNA fragments, the ability to non-enzymatically copy these molecules is essential. This process will not only permit an increase of the catalytic activity, but also of the RNA-pool size as well as the emergence of new catalysts because of copying errors. E) The RNA World. When the RNA pool contains molecules that are also catalysts of their own copying, as well as those of any other functional RNAs, a full-fledged RNA World is achieved.

processes must have intrinsically yielded RNA pools containing a large fraction of catalytically active molecules (by what is thus far an unknown process).

(ii) Environmental Conditions

Unfortunately, few clues about the original environmental conditions on primitive Earth remain due to the activity of the biosphere and plate tectonics. However, recent investigations suggest the presence of liquid water and continents as far back as 4.3 x 109 years ago. Higher volcanic activity and lower sunlight levels than today can also be expected. Thus life producing reactions could have occurred over a broad spectrum of conditions in terms of temperature, pressure, ionic strength, pH, etc. and have been located on land, deep within the Earth, in the oceans, or at interfaces between these regions.

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