A “cross-stitched” peptide with improved helicity and proteolytic stability

Abstract

A new computational approach to obtain quantitative energy profiles for helix folding was used in the design of orthogonal hydrocarbon and lactam bicyclic peptides. The proteolytically stable, "cross-stitched" peptide SRC2-BCP1 shows nanomolar affinity for estrogen receptor α and X-ray crystallography confirms a helical binding pose.

A Novel Long‐Range n to π* Interaction Secures the Smallest known α‐Helix in Water

“…B) Short‐range n→π* interactions (blue arrows) reported between backbone carbonyls in a protein α‐helix. C) The crystal structure (PDB ID: 5wgd) shows a peptide (green) bound to estrogen receptor α in an α‐helical conformation (yellow) that is constrained by a cyclic pentapeptide component, and key features include: i) a water molecule (red) hydrogen bonded to the amide linker (dash), ii) a long‐range n→π* interaction from the backbone CO to a side‐chain carbonyl carbon (red arrow), and iii) angles ( θ =97°, Θ =1.5°) and distance ( d =3.10 Å) indicating amide pyramidalization.…”

“…The root mean square fluctuation (RMSF) for 1 was less than for 2 , 3 or 5 in water, and less than for 1 in DMSO, which is consistent with structure stabilization by water solvation. A crystal structure for estrogen receptor α bound to a peptide containing the same Lys i to Asp i +4 linker as in 1 and 6 showed two peptides crystallized in the unit cell; an α‐helical one with a water H‐bonded to the linker amide C γ =Ο (Figure C), and a less helical one with no crystallized water (Figure S20). When the linker amide was water‐solvated (Figure C), it was pyramidalized with a long‐range n→π* interaction as in 6 but with a shorter distance d= 3.10 Å.…”

“…This side‐chain–side‐chain linker has been used in longer peptides to induce α‐helicity, which invariably increases affinity for proteins, chemical stability, and biological activity . Examples include hPTH, GLP‐1R, and ORL‐1 agonists, S. pneumoniae colony‐stimulating peptide CSP‐1, an RNA‐binding HIV‐1 Rev peptide, a Na + channel inhibitor, cJun‐binding cFos peptides, an estrogen receptor antagonist, and Bcl2 binding and MDM‐binding p53 peptides. Where helix structure in the cyclic peptide component was studied, the Lys i →Asp i +4 linkage was superior to similar linkers (i.e.…”

Macrocyclization of an all-D linear peptide improves target affinity and imparts cellular activity: A novel stapled α-helical peptide modality

“…GTP-ase [38], HIV-1 [39], Respiratory Syncytial Virus Entry [40,60], Ral GTP-ase [61], estrogen receptor-α [62] and BCL9 [63]. All these peptides exhibited increased helicity, increased proteolytic resistance and increased binding as compared to the corresponding single stapled peptides.…”

“…However the double-stapled peptide d PMI-δ(1-5,9-12) didn't have significant cellular activity with an EC50 of 34.6 µM, at 16 hrs [ Figure 8, Table 2]. Although recent studies have reported that the double-stapled peptides appear to follow the same trend as their single-stapled counterparts [ 38,39,40,[60][61][62], lack of cell activity observed for the double-stapled peptide here, demonstrate that enhanced cellular activity is not uniform. Although the molecular 13 mechanisms behind the increased cell permeability of the stitched D-peptide is unclear, it could be attributed to the increased conformational rigidity and/or increased hydrocarbon content of the peptide.…”