PMC

Structural differences of β-repeats. (A) Comparison of the PSAM structures. The structures are superimposed using the C-terminal globular domain. The N-terminal globular domain is omitted for clarity. The β-hairpin units in the PSAMs are labeled from the C terminus of the β-repeat segment to the N terminus starting from zero for the original β-hairpin unit. This “backward” direction is because a new copy of the β-hairpin unit is added N-terminal to the existing units. (B) The degrees of twist and bend between β-hairpin pairs in the PSAM crystal structures. β-hairpin pairs are denoted with the names of the two β-hairpin units connected with a hyphen. The definitions of twist, bend, and bend′ are provided in Materials and Methods.

Structures of β-repeat segments in PSAMs. (A) The backbone structure of the β-repeat segment in the crystal structure of OspA+3bh-sm1. The hydrogen bonds between backbone atoms are shown as dashed lines. (B) Side chain conformations of the β-repeat segment of OspA+3bh. Only the residues on the front face of the β-sheet are shown. The β-sheet backbone is shown as arrows. For clarity, the turn regions are omitted. The three cross-strand amino acid ladders are labeled with their respective amino acid compositions (“F/L,” “E/K,” and “T/I”). These ladders extend short and/or imperfect ones present in wild-type OspA (“FLFV,” “EKEK,” and “TVTI,” respectively) (). (C) Superposition of a total of 26 copies of the β-hairpin unit from the PSAM crystal structures. The backbone of the original β-hairpin unit from wild-type OspA is shown in green. Amino acid resides in the β-strand regions are labeled with an uppercase letter and residue number. Strand residues labeled in black have their side chain pointing toward the reader, and those labeled in gray have their side chain pointing away from the reader. Residues in the turn regions are labeled with a lowercase letter.

Demonstration of the propagation of small conformational differences of β-hairpin pairs (i.e., four-stranded building blocks) leading to substantial β-ribbon polymorphism. Larger peptide self-assemblies were modeled using six representative β-hairpin pairs. Different building blocks are shown in different colors (cyan, 5bh molecule C, β-hairpin units 4 and 3; blue, 5bh molecule A, units 3 and2; yellow, 5bh molecule A, units 5 and 4; green, 5bh molecule C, units 3 and 2; red, 2bh units 2 and 1), and only the backbone traces of the β-strand regions are shown for clarity. These β-hairpin pairs were superimposed using the first two β-strands (labeled with “1” and “2,” respectively). Different relative orientations of the third and fourth β-strands, with respect to the first and second, are evident. β-Ribbon superstructures shown at Right were constructed in a step-wise manner. Starting from a four-stranded building block, a copy of the building block was generated. The third and fourth β-strands of the original block and the first and second β-strands of the copy (which have the identical sequence and nearly identical conformation; C) were then superimposed. In this way, the third and fourth β-strands of the copy are now placed as the fifth and sixth β-strands of the original building block, and the relative orientation between adjacent two-stranded units (i.e., β-strands 1–2 and 3–4, and β-strands 3–4 and β-strand 5–6) is kept identical. These steps were iterated until a superstructure of sufficient length was generated.

Design principle, construction, and structures of peptide self-assembly mimics. (A) Scheme showing the concept of our “link-and-cap” strategy. A segment of peptide self-assembly is excised (step 1), covalently linked (step 2), and capped (step 3). (B–D) Construction of OspA-based PSAMs. (B) SDS/PAGE of purified PSAMs containing different numbers of additional hairpin units. (C) Correlation between the retention volume on a Superdex 75 column (Amersham; horizontal axis) and the expected size of the proteins. The data points are labeled with the numbers of hairpin inserts. (D) The free energy difference for the overall conformational stability as determined by chemical denaturation plotted as a function of the number of hairpin inserts. Stability measurements were performed as described in Yan et al. (). (E) Cartoon drawings of the atomic structures of PSAMs determined in this work. The N and C termini are designated in the OspA+1bh structure (far left). The β-hairpins in cyan are the original β-hairpin unit (strands 9 and 10 in wild-type OspA), those in red are additional copies, and those in blue are the homologous β-hairpin (strands 7 and 8). The N- and C-terminal globular domains are shown in gray.