X-ray Crystallographic Structure of a Teixobactin Derivative Reveals Amyloid-like Assembly

Abstract

This paper describes the X-ray crystallographic structure of a derivative of the antibiotic teixobactin and shows that its supramolecular assembly through the formation of antiparallel β-sheets creates binding sites for oxyanions. An active derivative of teixobactin containing lysine in place of allo-enduracididine assembles to form amyloid-like fibrils, which are observed through a thioflavin T fluorescence assay and by transmission electron microscopy. A homologue, bearing an N-methyl substituent, to attenuate fibril formation, and an iodine atom, to facilitate X-ray crystallographic phase determination, crystallizes as double helices of β-sheets that bind sulfate anions. β-Sheet dimers are key subunits of these assemblies, with the N-terminal methylammonium group of one monomer and the C-terminal macrocycle of the other monomer binding each anion. These observations suggest a working model for the mechanism of action of teixobactin, in which the antibiotic assembles and the assemblies bind lipid II and related bacterial cell wall precursors on the surface of Gram-positive bacteria.

Figure 1.

Teixobactin (1), Lys₁₀-teixobactin (2), and N-Me-d-Phe^I₁,N-Me-d-Gln₄Lys₁₀-teixobactin (3).

Figure 2.

(A) ThT fluorescence assay of Lys₁₀-teixobactin (2, four replicate runs with 120 μM peptide in PBS buffer at pH 7.4). (B) TEM images of the fibrils formed by Lys₁₀-teixobactin (2).

Figure 3.

X-ray crystallographic structure of a representative dimer of N-Me-d-Phe^I₁,N-Me-d-Gln₄,Lys₁₀-teixobactin (3). (A) Top view. (B) Side view.

Figure 4.

β-Sheet fibril formed by N-Me-d-Phe^I₁,N-Me-d-Gln₄Lys₁₀-teixobactin (3). (A) Top view. (B) Bottom view with hydrophobic side chains shown as spheres.

Figure 5.

Double helix of β-sheet fibrils formed by N-Me-d-Phe^I₁,N-Me-d-Gln₄,Lys₁₀-teixobactin (3). Sulfate anions are shown as spheres.

Figure 6.

Crystallographically based molecular model of an extended double helix of β-sheet fibrils formed by teixobactin analogue 2 and observed by TEM (Figure 2).