The Origins of Life
The complexity of modern biological life has long made it difficult to understand how life could emerge spontaneously from the chemistry of the early earth. The key to resolving this mystery lies in the simplicity of the earliest living cells. Through our efforts to synthesize extremely simple artificial cells, we hope to discover plausible pathways for the transition from chemical evolution to Darwinian evolution. We view the two key components of a primitive cell as a self-replicating nucleic acid genome, and a self-replicating boundary structure. We have uncovered simple and robust pathways for the coupled growth and division of primitive cell membranes, and have made significant experimental progress towards the synthesis of self-replicating nucleic acids. Recently, we have begun to investigate potential routes for the emergence of coded peptide synthesis from the chemistry of the RNA World. We are also interested in model systems that may provide a route to artificial life with a biochemistry that is distinct from that of existing biology.
References
Szostak JW. The Narrow Road to the Deep Past: In Search of the Chemistry of the Origin of Life. Angew. Chemie Int. Ed. Engl., 2017; 56:11037-11043.
Joyce GF, and Szostak JW. Protocells and RNA Self-Replication. Cold Spring Harbor Perspectives in Biology. 2018; 10(9). pii: a034801.
Nonenzymatic RNA Replication
The RNA genomes of the first cells are thought to have emerged from the nonenzymatic replication of short RNA strands. Several recent developments have enhanced our ability to copy RNA templates by primer extension. Through thermodynamic and kinetic studies, we demonstrated an important catalytic role for activated downstream nucleotides and oligonucleotides. We subsequently showed that this catalytic effect was due to the formation of a covalent imidazolium-bridged dinucleotide intermediate in primer extension. Mechanistic studies then led to the identification of 2-aminoimidazole as a superior nucleotide activating moiety. Our kinetic and crystallographic studies have provided insight into the mechanism of this key reaction, and to improvements in RNA copying chemistry that are both more prebiotically plausible and more accurate, efficient, and general.
Prebiotic chemistry is likely to have given rise not only to ribonucleotides, but also to related compounds with variations in the chemistry of the nucleobase and sugar components. Oligomers of such nucleotides would also have had significant heterogeneity in their backbone structure. Surprisingly, we find that ribonucleotides are typically more effective than likely alternatives in nonenzymatic template copying reactions. Our findings suggest a model for the transition from early heterogeneous nucleic acids to a more homogeneous form that is closer to modern RNA.
Going beyond simple template copying to multiple cycles of replication remains a key challenge. We have recently proposed the Virtual Circular Genome model for primordial RNA replication. This model avoids any requirement for specific primers, as well as problems related to the replication of the ends of linear sequences. We are currently working towards establishing a viable experimental system for nonenzymatic RNA replication based upon this model
References
Fahrenbach AF, Giurgiu C, Tam CP, Li L, Hongo Y, Aono M and Szostak JW. Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation. J. Am. Chem. Soc., 2017; 139:8780-8783.
Zhang W, Walton T, Li L, Szostak JW. Direct observation of nonenzymatic primer extension by X-ray crystallography. eLife, 2018; 7:e36422.
Kim S, Zhang W, O’Flaherty D, Zhou L, Rondo-Brevetta V, Szostak JW. A model for the emergence of RNA from a prebiotically plausible mixture of ribonucleotide arabinonucleotides and 2′-deoxynucleotides. J. Am. Chem. Soc., 2020; 142:2317-2326.
Zhou L, Ding D, Szostak JW. The virtual circular genome model for primordial RNA replication. RNA, 2021; 27:1-11.
Model Protocells
Very primitive cells may have consisted of a self-replicating nucleic acid genome, encapsulated within a self-replicating cell membrane. We have recently described robust pathways for the coupled growth and division of primitive cell membranes composed of fatty acids, which were likely to have been available prebiotically. Fatty acid membranes are remarkably permeable to charged small molecules such as nucleotides, which can be added to the outside of fatty acid vesicles so that RNA copying can proceed inside the vesicles. However, a major unresolved problem is that fatty acid vesicles are destroyed by the high concentrations of divalent cations such as Mg2+ that are required for RNA copying. This problem can be overcome by chelating the Mg2+ with citrate, but the search for a more plausible solution continues.
References
Budin I, Prywes N, Zhang N, and Szostak JW. Chain-length heterogeneity allows for the assembly of fatty acid vesicles in dilute solutions. Biophys. J., 2014; 107:1582-90.
O’Flaherty D, Kamat NP, Mirza FN, Li L, Prywes N, and Szostak JW. Copying of mixed sequence RNA templates inside model protocells. J. Am. Chem. Soc., 2018; 140:5171-5178.
Kindt J, Szostak JW, Wang A. Bulk self-assembly of giant, unilamellar vesicles. ACS Nano., 2020; 14:14627-14634.
The Origins of Translation
In modern biology, the ribosomal synthesis of proteins makes use of aminoacylated RNAs as substrates. Therefore, aminoacylated RNAs must have existed before the evolution of the primordial ribosome. However, the spontaneous chemical generation of such substrates is inefficient, and aminoacylated RNAs are unstable due to hydrolysis. We are searching for potential roles for aminoacylated RNAs that may have been important prior to the evolution of coded peptide synthesis. Recently we showed that aminoacylated RNAs can rapidly assemble into chimeric amino acid-bridged ribozymes that retain their native enzymatic activity. This potential role for RNA aminoacylation could have driven the evolution of aminoacyl-RNA synthetase ribozymes that were both sequence and amino acid specific, thereby setting the stage for the evolution of coded peptide synthesis.
References
Radakovic A, Wright T, Lelyveld V, Szostak JW. A Potential Role for Aminoacylation in Primordial RNA Copying Chemistry. Biochemistry, 2021; 60:477-488.
Radakovic A, DasGupta S, Wright TH, Aitken HRM, and Szostak JW. Nonenzymatic assembly of active chimeric ribozymes from aminoacylated RNA oligonucleotides. Proc. Natl. Acad. Sci. USA, 2022. Published online 2022 Feb 15; 119(7):e2116840119.