The key position: influence of staple location on constrained peptide conformation and binding

Abstract

Constrained α-helical peptides are showing potential as biological probes and therapeutic agents that target proteinprotein interactions. However, the factors that determine the optimal constraint locations are still largely unknown. Using the β-integrin/talin protein interaction as a model system, we examine the effect of constraint location on helical conformation, as well as binding affinity, using circular dichroism and NMR spectroscopy. Stapling increased the overall helical content of each integrin-based peptide tested. However, NMR analysis revealed that different regions within the peptide are stabilised, depending on constraint location, and that these differences correlate with the changes observed in talin binding mode and affinity. In addition, we show that examination of the atomic structure of the parent peptide provides insight into the appropriate placement of helical constraints.

Rational Design of a 310‐Helical PIP‐Box Mimetic Targeting PCNA, the Human Sliding Clamp

“…Linkage at this site would allow maintenance of native helical structure, while avoiding key PIP-box residues. Similar i to i+4 constraints have frequently been used to stabilize α-helical structure, [9,10] but 310 helices have generally been constrained by i, i+3 constraints, used in conjunction with the 310 helix promoting residue 2aminoisobutyric acid (Aib) (e.g., see [11][12][13] ). However, we reasoned that the particular orientation of this residue pair, combined with the natural 310 helical propensity of the p21 PIPbox sequence, would result in the desired structural stabilization.…”

“…[31] Peptide structures were calculated using a version of ARIA 2.3, modified to deal with lactam constrained peptides (Benjamin Bardiaux, Unité de Bioinformatique Structurale, Institut Pasteur). [10,14] Lactams were identified as being in the trans conformation by the presence of large NOEs between the lactam 'glutamate' side-chain protons to the lactam amide proton, as well as the absence of NOEs from the glutamate Hβs to the Hε protons of the lactam 'lysine' (ACR1), or Hγ protons of the lactam Dab residue (ACR2), so lactam φ-angles were set to -180°. Initial structures were calculated from NOE distance restraints and scalar coupling restraints.…”

Backbone thioamide directed macrocyclisation: lactam stapling of peptides

“…4–11 A wide range of peptide stapling strategies have been developed, including formation of disulfide bridges, 12 ring-closing metathesis, 13–16 CuAAC reactions, 17,18 and lactamisation. 19–23…”

A Bimane‐Based Peptide Staple for Combined Helical Induction and Fluorescent Imaging

“…A number of synthetic approaches have been developed to introduce such constraints, including lactam bridges, [6] all‐hydrocarbon staples, [5c,d,7] 1,2,3‐triazoles [8] and perfluoro aryl‐based linkers [9] . Each system has inherent benefits and limitations [2c,3a,7b,10] . A variety of dihalo reagents has also been used to define peptide backbone geometry by crosslinking cysteine side chains.…”

“…An auxiliary fluorescent tag can be attached to these peptidomimetics, and fluorescence can be monitored by confocal microscopy to allow imaging and tracking in a biological environment. However, introduction of a tag can adversely influence conformation, [15] cell permeability, [16] and the biological interaction under investigation [3a,17] . A significant advance would come with the development of a linker able to both conformationally constrain a peptide backbone, while also fluorescing to allow imaging and tracking.…”