Inhibition of Protein-Protein Interactions with α/β-Peptides

Targeting protein-protein interactions with α/β-peptides

Protein-protein interactions (PPI) regulate many essential biological processes through complementary binding sites on partner molecules. Aberrant PPIs can have severe implications for human health, so the ability to disrupt, restore, or otherwise monitor these recognition events is critical. The ability of peptides to present an array of side chains from a folded structure makes them prime candidates for targeting PPIs. However, peptides containing only traditional α-amino acids are subject to rapid proteolytic degradation, limiting their application in vivo without further modifications. Our lab seeks to overcome this disadvantage through the use of backbone-modified peptides, a class of molecules that have been shown to mimic the structure and function of natural counterparts while exhibiting greatly enhanced proteolytic stability.

α/β-Peptides derived from the Z-Domain scaffold

The “Z-domain” or “affibody” scaffold, derived from domain B of staphylococcal protein A, has been extensively studied for a wide range of applications, including tumor imaging, radiotherapy, and blocking PPI. The parent Z-domain (Z-IgG) is 58 residues in length and folds into a three-helix bundle. Combinatorial randomization of 13 residues on helices 1 and 2, followed by affinity-based selection, allows for the creation of Z-domain analogues that bind to a variety of therapeutically relevant targets, such as VEGF, TNFα, and HER2. Our lab combines two strategies to improve upon the synthetic accessibility and proteolytic stability of the original Z-domain scaffolds: (1) truncation of helix 3 followed by disulfide-crosslink stabilization of helices 1 and 2, and (2) insertion of unnatural residues on the now-vacant back faces of helices 1 and 2. This approach has been successfully employed to create α/β-peptides that bind to targets with affinity equal to their parent Z-domain miniproteins and are significantly enhanced proteolytic resistance. We hope to explore the therapeutic and diagnostic potential of these Z-domain α/β-peptides, as well as to extend this strategy to other Z-domain derivatives.

α/β-Peptides that mimic irregular secondary structure

The Gellman lab has been forefront at designing α/β-peptides that mimic α-helical secondary structure and modulate specific PPI. However, not all PPI interfaces are restricted to a single α-helical finger. Many recognition sites on proteins are comprised of a large and shallow surface area dominated by hydrophobic interactions. We use the irregular receptor-recognition site of VEGF as a model to develop α/β-peptides with complex secondary structure that retain affinity for VEGF. Previous efforts in our group have identified an α/β-peptide, HH4, that binds to VEGF with reasonable affinity and is more proteolytic stable than its α-amino acid parent peptide. Comparing the crystal structure of VEGF bound to HH4 to other published co-crystal structures of VEGF, we can rationally design a tighter binding α/β-peptide that resists proteolytic degradation. Using the results from this study, we hope to extend the design of α/β-peptides with irregular but discrete secondary structure to modulate different PPI.

Recent Publications:

6. "Iterative non-proteinogenic residue incorporation yields α/β-peptides with a helix-loop-helix tertiary structure and high affinity for VEGF." Checco, J.W.; Gellman, S.H. ChemBioChem. 2017, 18, 291-299.

5.  "α/β-Peptide Foldamers Targeting Intracellular Protein–Protein Interactions with Activity in Living Cells." Checco JW, Lee EF, Evangelista M, Sleebs NJ, Rogers K, Pettikiriarachchi A, Kershaw NJ, Eddinger GA, Belair DG, Wilson JL, Eller CH, Raines RT, Murphy WL, Smith BJ, Gellman SH, Fairlie D J. Am. Chem. Soc. 2015, 137, 11365.

4.“Residue-Based Preorganization of BH3-Derived α/β-Peptides: Modulating Affinity, Selectivity and Proteolytic Susceptibility in α-Helix Mimics.." Peterson-Kaufman KJ, Haase HS, Boersma MD, Lee EF, Fairlie WD, Gellman SH. ACS Chem. Biol. 2015, 10, 1667.

3. “Targeting diverse protein-protein interaction interfaces with α/β-peptides derived from the Z-domain scaffold.” Checco JW, Kreitler DF, Thomas NC, Belair DG, Rettko NJ, Murphy WL, Forest KT, Gellman SH. Proc. Natl. Acad. Sci. USA. 2015, 112, 4552.

2.“Consequences of periodic α-to-β3 residue replacement for immunological recognition of peptide epitopes.” Cheloha RW, Sullivan JA, Wang T, Sand JM, Sidney J, Sette A, Cook ME, Suresh M, Gellman SH. ACS Chem. Biol. 2015, 10, 844.

1. "Structure-Guided Rational Design of α/β-Peptide Foldamers with High Affinity for BCL-2 Family Prosurvival Proteins." Smith BJ, Lee EF, Checco JW, Evangelista M, Gellman SH, Fairlie WD. Chembiochem2013, 14, 1564.