Titratable Avidity Reduction Enhances Affinity Discrimination in Mammalian Cellular Selections of Yeast-Displayed Ligands.

摘要: Yeast surface display selections against mammalian cell monolayers have proven effective in isolating proteins with novel binding activity. Recent advances this technique allow for the recovery of clones even micromolar affinities. However, no efficient method has been shown affinity-based selection context. This study demonstrates effectiveness titratable avidity reduction using dithiothreitol to achieve goal. A series epidermal growth factor receptor fibronectin domains a range affinities are used quantitatively identify number ligands per yeast that yield strongest selectivity between strong, moderate, and weak Notably, ligand 3,000-6,000 2 nM binder yields 16-fold better than 17 binder. These lessons applied affinity maturation an EpCAM-binding population, yielding enriched pool significantly stronger analogous sorted by standard cellular methods. Collectively, offers facile approach yeast-displayed full-length targets generating promising further applications.