Screening of Yeast Display Libraries of Enzymatically Treated Peptides to Discover Macrocyclic Peptide Ligands

We present the construction and screening of yeast display libraries post-translationally modified peptides wherein site-selective enzymatic treatment linear is achieved using bacterial transglutaminase. To this end, we developed two alternative routes, namely (i) followed by with recombinant transglutaminase in solution; or (ii) intracellular co-expression to achieve peptide modification endoplasmic reticulum prior surface display. The efficiency was evaluated via orthogonal detection epitope tags integrated yeast-displayed flow cytometry, comparative cleavage putative cyclic vs. tobacco etch virus (TEV) protease. Subsequently, transglutaminase-treated were screened isolate binders N-terminal region Yes-Associated Protein (YAP) its WW domains magnetic selection fluorescence activated cell sorting (FACS). identified cyclo[E-LYLAYPAH-K] featured a KD 1.75 ?M for YAP 0.68 as well high binding selectivity against albumin lysozyme. These results demonstrate usefulness enzyme-mediated cyclization combinatorial identify binders.