Tracing the Origin of the Genetic Code and Thermostability to Dipeptide Sequences in Proteomes

The safekeeping of the genetic code has been entrusted to interactions between aminoacyl-tRNA synthetases and their cognate tRNA. In a previous phylogenomic study, chronologies of RNA substructures, protein domains and dipeptide sequences uncovered the early emergence of an 'operational' code in the acceptor arm of tRNA prior to the implementation of the 'standard' genetic code in the anticodon loop of the molecule. This history likely originated in peptide-synthesizing urzymes but was driven by episodes of molecular co-evolution and recruitment that promoted flexibility and protein folding. Here, we show that dipeptide sequences offer deep-time insights into the chronology of code emergence. A phylogeny describing the evolution of the repertoire of 400 canonical dipeptides reconstructed from an analysis of 4.3 billion dipeptide sequences across 1,561 proteomes revealed the overlapping temporal emergence of dipeptides containing Leu, Ser and Tyr, followed by those containing Val, Ile, Met, Lys, Pro, and Ala, all of which supported the operational RNA code. This strengthened a timeline of genetic code entry. The synchronous appearance of dipeptide-antidipeptide sequences along the dipeptide chronology supported an ancestral duality of bidirectional coding operating at the proteome level. Tracing determinants of thermal adaptation showed protein thermostability was a late evolutionary development and bolstered an origin of proteins in the mild environments typical of the Archaean eon. Our study uncovers a hidden evolutionary link between a protein code of dipeptides - arising from the structural demands of emerging proteins - and an early operational code shaped by co-evolution, editing, catalysis and specificity.