Peptide foldamer-based inhibitors of the SARS-CoV-2 S protein-human ACE2 interaction

Abstract

The entry of the SARS-CoV-2 virus into a human host cell begins with the interaction between the viral spike protein (S protein) and human angiotensin-converting enzyme 2 (hACE2). Therefore, a possible strategy for the treatment of this infection is based on inhibiting the interaction of the two abovementioned proteins. Compounds that bind to the SARS-CoV-2 S protein at the interface with the alpha-1/alpha-2 helices of ACE2 PD Subdomain I are of particular interest. We present a stepwise optimisation of helical peptide foldamers containing trans-2-aminocylopentanecarboxylic acid residues as the folding-inducing unit. Four rounds of optimisation led to the discovery of an 18-amino-acid peptide with high affinity for the SARS-CoV-2 S protein (K_d = 650 nM) that inhibits this protein-protein interaction with IC₅₀ = 1.3 µM. Circular dichroism and nuclear magnetic resonance studies indicated the helical conformation of this peptide in solution.

Keywords: BLI; COVID-19; foldamers; helix; peptides; protein-protein interaction.

Graphical abstract

Figure 1.

The strategy for the development of SARS-CoV-2 S protein-human ACE2 interaction inhibitors based on peptide foldamers.

Figure 2.

Crystal structure of LCB3-RBD complex (A), PDB id 7JZM (residues 1–18 of LCB3 are shown), and the modelled foldameric peptide 10 bound to the RBD of S protein (B). Peptides are shown in stick representation; trans-ACPC residues’ carbon atoms are coloured green, tryptophan residues’ carbon atoms are coloured pink. The surface of the RBD is coloured according to interpolated charge: blue – positive, grey – neutral, red – negative. Key intermolecular interactions are shown as dashed lines: green – hydrogen bonds, orange – charge assisted hydrogen bonds, pink – hydrophobic interactions, white – hydrogen bond donor/π interactions.

Figure 3.

CD spectra of (A) peptides (2–5) derived from LCB1 helix (1) and (B) peptides (6–10) derived from LCB3 helix (6), dissolved in potassium phosphate buffer, 50 mM, pH 7.5.

Figure 4.

BLI-based screening of the binding affinity properties of peptide 10 towards RBD. Association and dissociation steps are given for different concentrations of peptide 10, ranging from 1 to 100 µM.

Figure 5.

The modelled complex of peptide 12 with the RDB of the S protein. Peptide 12 is shown in stick representation; trans-ACPC residues’ carbon atoms are coloured green, mutated residues’ carbon atoms are coloured pink. The surface of the RBD is coloured according to interpolated charge: blue – positive, grey – neutral, red – negative. Key intermolecular interactions are shown as dashed lines: green – hydrogen bonds, orange – charge assisted hydrogen bonds, pink – hydrophobic interactions, white – hydrogen bond donor/π interactions.

Figure 6.

BLI-based screening of the binding affinity properties of peptide 12 towards RBD. Association and dissociation steps are given for the different concentrations of peptide 12, ranging from 1 to 100 µM.

Figure 7.

BLI-based screening of the binding affinity properties of peptide 27 towards RBD. Association and dissociation steps are given for different concentrations of peptide 27, ranging from 0.25 to 10 µM.

Figure 8.

Modelled complex of peptide 27 with the RBD of the S protein. Peptide 27 is shown in stick representation; trans-ACPC residues’ carbon atoms are coloured green, mutated residues’ carbon atoms are coloured pink. The surface of the RBD is coloured according to interpolated charge: blue – positive, grey – neutral, red – negative. Key intermolecular interactions are shown as dashed lines: green – hydrogen bonds, orange – charge assisted hydrogen bonds, pink – hydrophobic interactions, white – hydrogen bond donor/π interactions.

Figure 9.

CD spectra of peptides 33–41 dissolved in 50 mM potassium phosphate buffer, pH 7.5.

Figure 10.

BLI-based screening of the binding affinity properties of peptide 41 towards RBD. Association and dissociation steps are given for different concentrations of peptide 41, ranging from 0.25 to 10 µM.

Figure 11.

HTRF assay evaluation of the S RBD/ACE2 inhibitory activity of selected peptides.

Figure 12.

Regular NOESY contacts between HA and HN atoms of nonadjacent residues of peptide 27 (A). Contacts between i - i + 2, i - i + 3 and i - i + 4 are shown in green, blue and pink, respectively. Averaged structure of peptide 27 calculated on the basis of NMR-derived restraints shown in stick representation (B) and superimposition of 10 lowest energy structures (C). Green dotted lines represent hydrogen bonds.