Biology subjects

Aitken, H. R. M.

Publications and source records attributed to Aitken, H. R. M..

3 recordsLinked to original sources

bioRxiv2024

Overcoming nucleotide bias in the nonenzymatic copying of RNA templates.

The RNA World hypothesis posits that RNA was the molecule of both heredity and function during the emergence of life. This hypothesis implies that RNA templates can be copied, and ultimately replicated, without the catalytic aid of evolved enzymes. A major problem with nonenzymatic templated polymerization has been the very poor copying of sequences containing rA and rU. Here we overcome that problem by using a prebiotically plausible mixture of RNA mononucleotides and random-sequence oligonucleotides, all activated by methyl isocyanide chemistry, that direct the uniform copying of arbitrary-sequence templates, including those harboring rA and rU. We further show that the use of this mixture in copying reactions suppresses copying errors while also generating a more uniform distribution of mismatches than observed for simpler systems. We find that oligonucleotide competition for template binding sites, oligonucleotide ligation, and the template binding properties of reactant intermediates work together to reduce product sequence bias and errors. Finally, we show that iterative cycling of the activation chemistry and templated polymerization improves the yield of random-sequence products. These results for random-sequence template copying are a significant advance in the pursuit of nonenzymatic RNA replication.

molecular biology↗

bioRxiv2024

Structure-guided aminoacylation and assembly of chimeric RNAs

Coded ribosomal peptide synthesis could not have evolved unless its sequence and amino acid specific aminoacylated tRNA substrates already existed. We therefore wondered whether aminoacylated RNAs might have served some primordial function prior to their role in protein synthesis. Here we show that specific RNA sequences can be nonenzymatically aminoacylated and ligated to produce amino acid-bridged stem-loop RNAs. We used deep sequencing to identify RNAs that undergo highly efficient glycine aminoacylation followed by loop-closing ligation. The crystal structure of one such glycine-bridged RNA hairpin reveals a compact internally stabilized structure with the same eponymous T-loop architecture found in modern tRNA. We demonstrate that the T-loop assisted amino acid bridging of RNA oligonucleotides enables the rapid template-free assembly of a chimeric version of an aminoacyl-RNA synthetase ribozyme. We suggest that the primordial assembly of such chimeric ribozymes would have allowed the greater functionality of amino acids to contribute to enhanced ribozyme catalysis, providing a driving force for the evolution of sequence and amino acid specific aminoacyl-RNA synthetase enzymes prior to their role in protein synthesis.

biochemistry↗

bioRxiv2021

Non-enzymatic assembly of active chimeric ribozymes from aminoacylated RNA oligonucleotides

Aminoacylated tRNAs, which harbor a covalent linkage between amino acids and RNA, are a universally conserved feature of life. Because they are essential substrates for ribosomal translation, aminoacylated oligonucleotides must have been present in the RNA World prior to the evolution of the ribosome. One possibility we are exploring is that the aminoacyl ester linkage served another function before being recruited for ribosomal protein synthesis. The nonenzymatic assembly of ribozymes from short RNA oligomers under realistic conditions remains a key challenge in demonstrating a plausible pathway from prebiotic chemistry to the RNA World. Here, we show that aminoacylated RNAs can undergo template-directed assembly into chimeric amino acid-RNA polymers that are active ribozymes. We demonstrate that such chimeric polymers can retain the enzymatic function of their all-RNA counterparts by generating chimeric hammerhead, RNA ligase, and aminoacyl transferase ribozymes. Amino acids with diverse side chains form linkages that are well tolerated within the RNA backbone, potentially bringing novel functionalities to ribozyme catalysis. Our work suggests that aminoacylation chemistry may have played a role in primordial ribozyme assembly. Increasing the efficiency of this process provides an evolutionary rationale for the emergence of sequence and amino acid specific aminoacyl-RNA synthetase ribozymes, which could then have generated the substrates for ribosomal protein synthesis. Significance StatementThe emergence of the primordial ribosome from the RNA World would have required access to aminoacylated RNA substrates. The spontaneous generation of such substrates without enzymes is inefficient, and it remains unclear how they could be selected for in a prebiotic milieu. In our study we identify a role for aminoacylated RNA in ribozyme assembly, a longstanding problem in the origin of life research. We show that aminoacylated RNAs, but not unmodified RNAs, rapidly assemble into chimeric amino acid-bridged ribozymes that retain their native enzymatic activity. Our work therefore addresses two key challenges within the origin-of-life field: we demonstrate assembly of functional ribozymes and we identify a potential evolutionary benefit for RNA aminoacylation that is independent of coded peptide translation.

biochemistry↗