Toward the development of peptide nanofilaments and nanoropes as smart materials

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   Fig. 1.

   Protein design of a phased-hydrophobic polymer. (A) Surface representation of CpA, color-coded in green to highlight the hydrophobic residues. The amino acid sequence represents a tandem repeat of the first two heptads from the coiled-coil domain from the yeast transcription factor, GCN4, with a two-alanine insertion between the two pairs of heptads. The sequence of the designed peptide is shown in juxtaposition to the native sequence from GCN4 to highlight the differences. The methionine at the first position has been mutated to cysteine to allow crosslinking of the polymers through the formation of disulfide bonds. (B) Molecular model of the CpA dimer interface, showing the packing interactions between residues highlighted as space-filling representations. A staggered arrangement of individual helix interactions fosters polymerization as each unsatisfied hydrophobic surface on the growing chain recruits additional peptide units, as shown by the cylinders below the model. (C) Molecular model of a self-assembled polymer, illustrating several higher-order structural features. The two-alanine insertion results in a phase shift of ≈200° of the C-terminal hydrophobic surface, relative to the N-terminal hydrophobic surface, leading to a third level of supercoiling. Rotation about an axis perpendicular to the direction of coiling allows for a view down the supercoil, allowing for a visual representation of the width (≈6.7 nm).

   
   
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   Fig. 2.

   Tapping-mode AFM images reveal filamentous polymers and fibrils (nanoropes). Samples were prepared by using 144 μM peptide and 10 mM Tris·HCl, pH 8.0. (a) Phase image of nanofilaments formed in 1.5 M NaCl. (b) Topography image of nanofilaments formed in 0.75 M (NH₄)₂SO₄; z-scale, 25 nm. The nanofilaments are much longer than those in a and show greater tendency to associate. (c) Phase image showing the same area and magnification as in b. The internal structure is better resolved in the phase images.

   
   
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   Fig. 3.

   The stability of the polymers can be modulated by environmental conditions. (A) Spectra were measured at 25°C by using 102 μM peptide and 10 mM Tris·HCl, pH 8.0. (B) Polymerizing systems under thermodynamic control depend on monomer (or peptide) concentration. Samples were measured at 25°C and contained 2 M NaCl and 10 mM Tris·HCl, pH 8.0. (C) Comparison of salt effects on helix stability suggests sulfate preferentially stabilizes structure. Samples were measured at 25°C by using 144 μM peptide and 10 mM Tris·HCl, pH 8.0. NaCl (▪), (NH₄)₂SO₄ (▴), and Na₂SO₄ (•). (Inset) The effect of glycerol on helix content is shown. (D) Temperature unfolding experiments reveal the stability of the CpA polymers as a function of NaCl concentration. Samples were measured by using 144 μM peptide and 10 mM Tris·HCl, pH 8.0. At the highest NaCl concentration, a single transition is seen whose midpoint is ≈55°C. (Inset) The helical spectrum before (solid line) and after (dashed line) thermal denaturation, demonstrating quantitative reversibility, is shown.