Automated Flow Synthesis of Combinatorial Protein-Length Polyamide Libraries

Flow synthesis with building block mixtures permits the generation of hypervariable polyamide libraries with quintillions of members on a timescale of < 1 hour per library. The semi-automated flow platform on which we have previously developed this technique, however, lacks the efficiency required to make protein-length polyamide chains. To facilitate the direct engraftment of variable regions into longer polyamide chains, we adapted mixture-based library synthesis to automated flow peptide synthesis (AFPS) systems previously optimized for the production of 100-200-mer synthetic proteins. Using this approach, we demonstrate the single-shot synthesis of a 101-mer library with content approaching quintillions of members consisting of both L- and D- amino acid building blocks. AFPS thus permits the incorporation of hypervariable polyamide libraries into synthetic protein-length polyamide scaffolds.