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bioRxiv · 10.1101/2020.08.13.249722

Use of target-displaying magnetized yeast in screening mRNA display peptide libraries to identify ligands

Bowen, J. D.·Bacon, K. B.·Reese, H. R.·Rao, B. M.·Menegatti, S.
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Abstract

This work presents the first use of yeast-displayed protein targets for screening mRNA display libraries of cyclic and linear peptides. The WW domains of Yes-Associated Protein 1 (WW-YAP) and mitochondrial import receptor subunit TOM22 were adopted as protein targets. Yeast cells displaying WW-YAP or TOM22 were magnetized with iron oxide nanoparticles to enable the isolation of target-binding mRNA-peptide fusions. Equilibrium adsorption studies were conducted to estimate the binding affinity (KD) of selected WW-YAP-binding peptides: KD values of 37 M and 4 M were obtained for cyclo[M-AFRLC-K] and its linear cognate, and 40 M and 3 M for cyclo[M-LDFVNHRSRG-K] and its linear cognate, respectively. TOM22-binding peptide cyclo[M-PELNRAI-K] was conjugated to magnetic beads and incubated with yeast cells expressing TOM22 and luciferase. A luciferase-based assay showed a 4.5-fold higher binding of TOM22+ yeast compared to control cells. This work demonstrates that integrating mRNA and yeast display accelerates the discovery of biospecific peptides.

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Bowen, J. D., Bacon, K. B., Reese, H. R., Rao, B. M., Menegatti, S.. 2020-08-14. Use of target-displaying magnetized yeast in screening mRNA display peptide libraries to identify ligands. https://doi.org/10.1101/2020.08.13.249722

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