(272e) Constructing, Evaluating, and Screening Bioconjugates on the Yeast Surface

The introduction of non-canonical amino acids (ncAAs) into proteins offers attractive opportunities for engineering new classes of reagents, diagnostics, and therapeutics. We present here a ncAA-compatible yeast surface display platform that enables the construction, evaluation, and screening of bioconjugates on the yeast surface. This work utilizes azide- and ketone-containing antibody constant regions (Fcs) and antigen-binding Fcs (Fcabs) to demonstrate several important aspects of the platform. Binding experiments indicate that ncAA-substituted constructs remain well-folded and, in the case of Fcabs, able to bind to target antigens. Moreover, azide-containing constructs can be covalently modified on the yeast surface under mild conditions while retaining their thermal stability. Importantly, model sorting experiments lead to the efficient enrichment of constructs based on either chemical modification status or antigen binding. Cells displaying azide-containing Fcs can be selectively modified and enriched from an excess of cells displaying Fcs containing only canonical amino acids. Mixtures of successfully constructed conjugates in which only a small portion recognize a target antigen can be enriched for the conjugates exhibiting antigen recognition. The approaches presented here will enable high throughput characterization and screening of libraries of protein-small molecule hybrids for the discovery of therapeutic leads and numerous other biological applications.