Binary combinatorial scanning reveals potent poly-alanine-substituted inhibitors of protein-protein interactions

Xiyun Ye; Yen-Chun Lee; Zachary P Gates; Yingjie Ling; Jennifer C Mortensen; Fan-Shen Yang; Yu-Shan Lin; Bradley L Pentelute

doi:10.1038/s42004-022-00737-w

Binary combinatorial scanning reveals potent poly-alanine-substituted inhibitors of protein-protein interactions

Commun Chem. 2022 Oct 14;5(1):128. doi: 10.1038/s42004-022-00737-w.

Authors

Xiyun Ye^#¹, Yen-Chun Lee^#^{1

2}, Zachary P Gates^{1

3

4}, Yingjie Ling⁵, Jennifer C Mortensen⁵, Fan-Shen Yang⁶, Yu-Shan Lin⁵, Bradley L Pentelute^{7

8

9

10}

Affiliations

¹ Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA, 02139, USA.
² Department of Chemistry, National Cheng Kung University, No.1, University Road, Tainan City, 701, Taiwan.
³ Institute of Sustainability for Chemicals, Energy and Environment (ISCE2), Agency for Science, Technology and Research (A*STAR), 8A Biomedical Grove, Singapore, 138665, Singapore.
⁴ Disease Intervention Technology Laboratory (DITL), Agency for Science, Technology and Research (A*STAR), 61 Biopolis Drive, Singapore, 138673, Singapore.
⁵ Department of Chemistry, Tufts University, 62 Talbot Avenue, Medford, MA, 02155, USA.
⁶ Department of Chemistry and Frontier Research Center on Fundamental and Applied Sciences and Matters, National Tsing Hua University, 101, Sec. 2, Guang-Fu Road, Hsinchu, 300, Taiwan.
⁷ Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA, 02139, USA. blp@mit.edu.
⁸ The Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, 500 Main Street, Cambridge, MA, 02142, USA. blp@mit.edu.
⁹ Center for Environmental Health Sciences, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA, 02139, USA. blp@mit.edu.
¹⁰ Broad Institute of MIT and Harvard, 415 Main Street, Cambridge, MA, 02142, USA. blp@mit.edu.

^# Contributed equally.

Abstract

Establishing structure-activity relationships is crucial to understand and optimize the activity of peptide-based inhibitors of protein-protein interactions. Single alanine substitutions provide limited information on the residues that tolerate simultaneous modifications with retention of biological activity. To guide optimization of peptide binders, we use combinatorial peptide libraries of over 4,000 variants-in which each position is varied with either the wild-type residue or alanine-with a label-free affinity selection platform to study protein-ligand interactions. Applying this platform to a peptide binder to the oncogenic protein MDM2, several multi-alanine-substituted analogs with picomolar binding affinity were discovered. We reveal a non-additive substitution pattern in the selected sequences. The alanine substitution tolerances for peptide ligands of the 12ca5 antibody and 14-3-3 regulatory protein are also characterized, demonstrating the general applicability of this new platform. We envision that binary combinatorial alanine scanning will be a powerful tool for investigating structure-activity relationships.

Abstract

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