Structured Cyclic Peptides That Bind the EH Domain of EHD1

Abstract

EHD1 mediates long-loop recycling of many receptors by forming signaling complexes using its EH domain. We report the design and optimization of cyclic peptides as ligands for the EH domain of EHD1. We demonstrate that the improved affinity from cyclization allows fluorescence-based screening applications for EH domain inhibitors. The cyclic peptide is also unusually well-structured in aqueous solution, as demonstrated using nuclear magnetic resonance-based structural models. Because few EH domain inhibitors have been described, these more potent inhibitors will improve our understanding of the roles of EHD1 in the context of cancer invasion and metastasis.

Conformational Restriction of Peptides Using Dithiol Bis-Alkylation

“…This makes it very challenging to predictively design specific cross-links to stabilize a desired loop structure. Currently, identifying the proper linker chemistry, length, and positioning can only be done in an iterative process (Berthelot, Gonçalves, Laïn, Estieu-Gionnet, & Déléris, 2006; Cudic, Wade, & Otvos, 2000; Hayouka et al, 2012; Kamens, Eisert, Corlin, Baleja, & Kritzer, 2014; Qvit et al, 2009). One way to accelerate, this process is to introduce diverse conformational constraints at a late stage of synthesis.…”

Eps15 Homology Domain-containing Protein 3 Regulates Cardiac T-type Ca2+ Channel Targeting and Function in the Atria

“…This approach may offer a way to specifically tune ion channel trafficking to tune cardiac excitability in disease. Indeed, this potential has sparked the active development of EHD inhibitors (63,64). We have previously demonstrated that in vitro EHD3 overexpression is sufficient to tune cell excitability (7), suggesting that targeting endosomal transport may be a feasible approach.…”

Molecular dynamics simulation of the interactions between EHD1 EH domain and multiple peptides

“…Studies have shown that cyclic NPF-containing peptides bind to the N-terminal EH domain with higher affinities than the linear ones (Yamabhai et al, 1998;de Beer et al, 2000). And a new cyclic peptide designed for the EHD1 EH domain has obtained nearly 4-fold improvement in affinity in contrast to a typical linear peptide (Kamens et al, 2014). Usually, the β-turn conformation that is adopted by the NPF motif of bound state was well stabilized in these cyclic peptides.…”

“…However, just as in the observation of Kim et al (2001), the rigid cyclic peptide may induce larger conformational changes of its protein partner, slow the association rate, and thus get a lower affinity. Weaker binding affinities were also observed when increasing or decreasing the ring size of a good cyclic peptide (Kamens et al, 2014). Therefore, how to design a specific cyclic peptide with an appropriate conformation is a big challenge for the future.…”

“…EHD1 and other proteins involved in the endocytic events are thus becoming potential cancer targets for peptide inhibitor design. Recently, several cyclic peptides have been designed with higher affinities toward the EHD1 EH domain (Kamens et al, 2014).…”