Amide bond formation is one of the most important reactions in both chemistry and biology 1-4 , but there is currently no chemical method to achieve α-peptide ligation in water that tolerates all twenty proteinogenic amino acids at the peptide ligation site. The universal genetic code establishes the biological role of peptides predates Life's last universal common ancestor and that peptides played an essential role in the origins of Life 5-9 . The essential role of sulfur in the citric acid cycle, non-ribosomal peptide synthesis and polyketide biosynthesis points towards thioester-dependent peptide ligations preceding RNA-dependent protein synthesis during the evolution of Life 5,9-13 . However, a robust mechanism for aminoacyl thioester formation has never been demonstrated 13 . Here, we report a chemoselective, high yielding a-aminonitrile ligation that exploits only prebiotically plausible molecules-hydrogen sulfide, thioacetate 12,14 and ferricyanide 12,14-17 or cyanoacetylene 8,14 -to yield apeptides in water. The ligation is extremely selective for a-aminonitrile coupling and tolerates all 20 proteinogenic amino acid residues. Two essential features enable the peptide ligation in water: 1) the reactivity and pKaH of a-aminonitriles makes them compatible with ligation at neutral pH, and 2) Nacylation stabilises the peptide product and activates the peptide precursor to (biomimetic) N®C peptide ligation. Our model unites prebiotic aminonitrile synthesis and biological a-peptides, suggesting short N-acyl peptide nitriles were plausible substrates during early evolution.To improve the efficiency and selectivity of peptide ligation in water we sought to develop a novel mechanism for non-enzymatic peptide synthesis, which would operate via biomimetic N®C ligation in near-neutral pH water, and we suspected that a combination of sulfur and nitrile chemistry would be required ( Fig. 1a) 8,9,14,[18][19][20][21] . Proteinogenic a-aminonitriles (AA-CN) are readily synthesised 8,18 , and their direct ligation would provide the simplest prebiotic pathway to peptides. Unfortunately, incubation of AA-CN in water results in extremely ineffective peptide synthesis 22 . a-Amino acids (AA) are widely assumed to be prebiotic precursors of peptides, but the harsh conditions (typically strongly acidic or alkaline solutions) required for AA formation from AA-CN are incompatible with the integrity of both peptides and electrophilic activating agents. Therefore, we sought a more congruent and direct pathway from a-aminonitriles to a-peptides, and although the conversion of AA-CN to AA-SH has never been reported 23 , harnessing the AA-CN nitrile moiety for thioacid synthesis seemed more prudent than dissipating its activation through exhaustive hydrolysis.Orgel has previously suggested that a-aminothioacids (AA-SH) 16 might offer an interesting alternative to biological thioesters 10,11 . AA-SH unite excellent aqueous stability with highly selective (electrophilic or oxidative) activation 12,14,16,24 , but their prebiotic synthesis...

It has recently emerged that the succinimide linkage of a maleimide thiol addition product is fragile, which is a major issue in fields where thiol functionalisation needs to be robust. Herein we deliver a strategy that generates selective cysteine thiol labelling reagents, which are stable to hydrolysis and thiol exchange.

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