Helix formation and capping energetics of arginine analogs with varying side chain length

Abstract

Arginine (Arg) has been used for recognizing negatively charged biological molecules, cell penetration, and oligosaccharide mass signal enhancement. The versatility of Arg has inspired the need to develop Arg analogs and to research the structural effects of incorporating Arg analogs. Accordingly, we investigated the effect of Arg side chain length on helix formation by studying 12 Ala-based peptides containing the Arg analogs (S)-2-amino-6-guanidino-hexanoic acid (Agh), (S)-2-amino-4-guanidinobutyric acid (Agb), and (S)-2-amino-3-guanidinopropionic acid (Agp). Solid phase guanidinylation with orthogonal protection strategies was necessary to synthesize Agb- and Agp-containing peptides using Fmoc-based chemistry. The fraction helix for the peptides was determined by circular dichroism spectroscopy, and used to derive the statistical mechanical parameters and energetics for N-capping, C-capping, and helix propagation (propensity). All four Arg analogs were unfavorable for N-capping. The C-cap parameter followed the trend Agp<Agb<Arg<Agh, showing more favorable C-cap energetics with increasing side chain length. In contrast, helix propensity followed the trend Agp<Agb<Arg>Agh, highlighting the uniqueness of the Arg side chain length in helix formation. Molecular mechanics calculations and a survey on protein structures were consistent with the experimental results. Furthermore, calculations and survey both showed that the g- conformation for the χ1 dihedral was present for the first two residues at the N-terminus of helices, but not favored in the center or C-terminus of helices due to sterics. These results should serve as the foundation for developing Arg-related bioactive compounds and technologies.

Effects of Arginine Deimination and Citrulline Side‐Chain Length on Peptide Secondary Structure Formation

“…The helix propensity w for the Cit analogues followed the trend Aup < Aub < Cit > Auh (Table ). This trend was similar to the helix propensity trend for Arg analogues (Agp < Agb < Arg > Agh) . The helix propensity increased with increasing side‐chain length up to three methylene units (Cit and Arg) in both trends.…”

“…The α‐helical peptides were designed according to the Ala‐based peptides studied by Baldwin and later by our group (Table ) . Tyrosine was incorporated in order to facilitate concentration determination by UV/Vis absorption spectroscopy .…”

“…The CD spectra for the α‐helical peptides were acquired in 1 m m borate, 1 m m citrate, 1 m m phosphate with 1 m NaCl at pH 7 (Figure ) . The peptides bear multiple positive charges at pH 7 and are not amphiphilic, so these peptides should not associate in neutral aqueous buffers.…”

“…The CD signal at 222 nm for the peptides were used to derive the fraction helix (Table S1), which was used to derive the statistical mechanical helix propensity ( w ) for each residue at the guest position, by using modified Lifson–Roig theory by the least‐squares method (Table ) . The helix propensities obtained by the least‐squares method provide the optimum values to fit the data for both KXaa9 and KXaa14 peptides for a given guest residue (Xaa).…”

“…Deriving helix propensity : The fraction helix ( f helix ) of each peptide was derived from the CD data at 222 nm by f helix =([ θ ] 222 /(40 000(1–2.5/ N ))) in which N is the number of backbone amide units . The helix propensity ( w ) for a given residue at the guest position (Xaa) was derived from f helix by using modified Lifson–Roig theory by the least squares method to account for both KXaa9 and KXaa14 peptides. The helix formation energetics (Δ G helix ) for a given residue Xaa were derived from the corresponding helix propensity ( w ) by Δ G helix =− RT ln w .…”