Indirect Formation\nof Peptide Bonds as a Prelude to\nRibosomal Transpeptidation

The catalytic competency of the ribosome in extant protein\nbiosynthesis\nis thought to arise primarily from two sources: an ability to precisely\njuxtapose the termini of two key substrates3′-aminoacyl\nand <i>N</i>-acyl-aminoacyl tRNAsand an ability\nto ease direct transpeptidation by their desolvation and encapsulation.\nIn the absence of ribosomal, or enzymatic, protection, however, these\nactivated alkyl esters undergo efficient hydrolysis, while significant\nentropic barriers serve to hamper their intermolecular cross-aminolysis\nin bulk water. Given that the spontaneous emergence of a catalyst\nof comparable size and sophistication to the ribosome in a prebiotic\nRNA world would appear implausible, it is thus natural to ask how\nappreciable peptide formation could have occurred with such substrates\nin bulk water without the aid of advanced ribozymatic catalysis. Using\na combination of fluorine-tagged aminoacyl adenylate esters, in situ\nmonitoring by <sup>19</sup>F{<sup>1</sup>H} NMR spectroscopy, analytical\ndeconvolution of kinetics, pH–rate profile analysis, and temperature-dependence\nstudies, we here explore the mechanistic landscape of indirect amidation,\nvia transesterification and O-to-N rearrangement, as a highly efficient,\nalternative manifold for transpeptidation that may have served as\na prelude to ribosomal peptide synthesis. Our results suggest a potentially\noverlooked role for those amino acids implicated by the cyanosulfidic\nreaction network with hydroxyl side chains (Ser and Thr), and they\nalso help to resolve some outstanding ambiguities in the broader literature\nregarding studies of similar systems (e.g., aminolyzes with Tris buffer).\nThe evolutionary implications of this mode of peptide synthesis and\nthe involvement of a very specific subset of amino acids are discussed.