Self-assembly of the ionic peptide EAK16: the effect of charge distributions on self-assembly

Abstract

Amphiphilic peptides suspended in aqueous solution display a rich set of aggregation behavior. Molecular-level studies of relatively simple amphiphilic molecules under controlled conditions are an essential step toward a better understanding of self-assembly phenomena of naturally occurring peptides/proteins. Here, we study the influence of molecular architecture and interactions on the self-assembly of model peptides (EAK16s), using both experimental and theoretical approaches. Three different types of EAK16 were studied: EAK16-I, -II, and -IV, which have the same amino acid composition but different amino acid sequences. Atomic force microscopy confirms that EAK16-I and -II form fibrillar assemblies, whereas EAK16-IV forms globular structures. The Fourier transform infrared spectrum of EAK16-IV indicates the possible formation of a beta-turn structure, which is not found in EAK16-I and -II. Our theoretical and numerical studies suggest the underlying mechanism behind these observations. We show that the hairpin structure is energetically stable for EAK16-IV, whereas the chain entropy of EAK16-I and -II favors relatively stretched conformations. Our combined experimental and theoretical approaches provide a clear picture of the interplay between single-chain properties, as determined by peptide sequences (or charge distributions), and the emerging structure at the nano (or more coarse-grained) level.

FIGURE 1

Schematic three-dimensional molecular model drawn with ChemSketch, based on energy minimization: (a) EAK16-II and (b) EAK16-IV. Carbon atoms are cyan, oxygen atoms are red, nitrogen atoms are blue, and hydrogen atoms are white. In this conformation, all of the hydrophobic alanine side chains face in one direction, and all of the lysine and glutamic acid side chains face in the other direction to create two distinct faces. On the polar face, glutamic acid alternates with lysine. Below the molecular models are simplified representations of the peptides with the individual amino acids shown as spherical monomers.

FIGURE 2

(a) Mapping onto an effective bending energy. A stretched chain can be transformed into a hairpin by successive rotations of dihedral angles φ_i and φ_i+1. However, one can achieve a similar conformational change through a rotation of bond angle θ. This implies that we can represent the torsional energy of freely rotating chain model in terms of an effective bending energy to a certain extent. (b) Polymer model used in the simulations. Different symbols represent different charged monomers (shaded circles, neutral; solid diamonds, negatively charged; open diamonds, positively charged monomers). See Computer Simulations for detail.

FIGURE 3

AFM images of EAK16-I (a), -II (b), and -IV (c) at a concentration of 0.1 mg/mL. EAK16-I and -II form fibrillar nanostructures, whereas EAK16-IV forms globular ones. The scan size of the images is 2 × 2 μm² and z-scale is 5 nm.

FIGURE 4

(a) FT-IR spectra of EAK16-I, -II, and -IV for peptide concentrations 1.0, 1.0, 3.0 mg/mL, respectively. Distinct from the other two peptides, EAK16-IV has a broad peak centered at ∼1675 cm⁻¹. This particular peak is attributed to the formation of turn structure. (b) FT-IR of EAK16-II at c = 0.08 mg/mL.

FIGURE 5

(a) Energies in conformational changes from stretched to folded EAK16-II and -IV. For II has to overcome an energy barrier of 1.6 k_BT, whereas IV loses 2.4 k_BT. Thus, energetically, II prefers to be extended whereas IV prefers to fold. (b) The distribution of the end-to-end distance P(R, L) = 4πR²G(R, L) of CWLC I, II, and IV. The asymmetry in the distribution shows that EAKs with a relatively flexible backbone usually look like “globules” in solution, conformations between fully stretched and completely folded ones. The chain entropy favors this conformation over the other cases, because it has more conformational degrees of freedom (hence more entropy). Each curve was constructed from ∼43 million MC steps. Note how the charge polarities of chain can affect P(R, L) and its peak position. Here, = 2 aa, κ = 0, and σ = 0.9 aa. See also Appendix III.

FIGURE 6

Cloud-map representation of the distribution of the end-to-end distance R. The clouds describe a uniform sampling of an energy space by a one-dimensional random walk, i.e., the distribution of dots represents the distribution of R (see text for the detail). The color gradients represent the “temperature” T in the multicanonical sampling. In other words, the distribution of R at a given temperature is described by dots in the same color. Red/blue dots correspond to low/high T, at which the chain conformation is determined mainly by energy entropy. The minimum-energy conformation for type-I is a “worm” whereas, for type-IV, it is a slightly crossed hairpin. (Left) Type-II sequence. The two chains shown below the cloud map are MC generated, typical conformations in the red regions. (Top right) Type-I. (Bottom right) Type-IV.

FIGURE 7

AFM images of EAK16-IV aggregation. Morphology changes from globular to fibrillar in type-IV peptides. (b) c = 3 mg/mL (d_inter = 9.7 nm). (a) c = 7 mg/mL (d_inter = 7.3 nm). The scan size of the images is 1.5 × 1.5 μm² and z-scale is 6 nm.

FIGURE 8

Two-chain electrostatic interaction energy Φ versus center-to-center distance d_c2c between chains, in units of k_BT and aa, respectively. Three types of charged WLC, EAK16-I, -II, and -IV, were examined, and ∼0.2 billion MC steps and 20,000 chain conformations were sampled for each curve. Note that the interchain interaction is almost two orders of magnitude smaller than the thermal energy k_BT. On the other hand, EAK16-I almost behaves like an uncharged peptide.

FIGURE 9

AFM images of EAK16-II aggregation. (a) c = 0.3 mg/mL. (b) c = 0.08 mg/mL. The scan size of the images is 2 × 2 μm² and z-scale is 2 nm (a) and 7 nm (b).