Relationship between side-chain branching and stoichiometry in β(3)-peptide bundles

Abstract

The stability and stoichiometry of β(3)-peptide bundles is influenced by side-chain identity. β(3)-peptides containing β(3)-homoleucine on one helical face assemble into octamers, whereas those containing β(3)-homovaline form tetramers. From a structural perspective, the side chains of β(3)-homoleucine and β(3)-homovaline differ in terms of both side-chain length and γ-carbon branching. To evaluate the extent to which these two parameters control β(3)-peptide bundle stoichiometry, we synthesized the β(3)-peptide Acid-3Y, which contains β(3)-homoisoleucine in place of β(3)-homoleucine or β(3)-homovaline. Acid-3Y assembles into a stable tetramer whose stability resembles that of the previously characterized Acid-VY tetramer. These results suggest that β(3)-peptide bundle stoichiometry is dominated by the presence or absence of γ-carbon branching on core side chains.

Fig. 1

(a) Helical net and linear representations of β³-peptides discussed in this work and (b) comparison between β³-hLeu, β³-hVal and β³-hIle side chains. Letters represent standard amino acid abbreviations; O = ornithine.

Fig. 2

Self-association of Acid-3Y monitored by sedimentation equilibrium analytical ultracentrifugation (SE-AU) and fit to a monomer-n-mer equilibrium model. Results are shown for [Acid-3Y] = 27.5 µM. Data points are represented as open circles with colors corresponding to the indicated speeds. Black lines indicate fits to (a) n = 4; (b) n = 3; (c) n = 5; and (d) n = 8 models.

Fig. 3

Biophysical characterization of the Acid-3Y bundle. (a) Wavelength-dependent circular dichroism (CD) spectra of Acid-3Y at concentrations between 20 and 100 µM; (B) Plot of MRE₂₁₂ versus [Acid-3Y] fit to an ideal monomer-tetramer equilibrium (red line); (C) Plot of δMRE/δT as a function of T for five different concentrations of Acid-3Y; (D) Ratio of ANS fluorescence (I/I₀) in the presence of the indicated concentration of Acid-1Y (octamer), Acid-VY (tetramer) and Acid-3Y (tetramer). The concentration of ANS was 10 µM.