Coiled coils at the edge of configurational heterogeneity. Structural analyses of parallel and antiparallel homotetrameric coiled coils reveal configurational sensitivity to a single solvent-exposed amino acid substitution - PubMed (original) (raw)

Coiled coils at the edge of configurational heterogeneity. Structural analyses of parallel and antiparallel homotetrameric coiled coils reveal configurational sensitivity to a single solvent-exposed amino acid substitution

Maneesh K Yadav et al. Biochemistry. 2006.

Abstract

A detailed understanding of the mechanisms by which particular amino acid sequences can give rise to more than one folded structure, such as for proteins that undergo large conformational changes or misfolding, is a long-standing objective of protein chemistry. Here, we describe the crystal structures of a single coiled-coil peptide in distinct parallel and antiparallel tetrameric configurations and further describe the parallel or antiparallel crystal structures of several related peptide sequences; the antiparallel tetrameric assemblies represent the first crystal structures of GCN4-derived peptides exhibiting such a configuration. Intriguingly, substitution of a single solvent-exposed residue enabled the parallel coiled-coil tetramer GCN4-pLI to populate the antiparallel configuration, suggesting that the two configurations are close enough in energy for subtle sequence changes to have important structural consequences. We present a structural analysis of the small changes to the helix register and side-chain conformations that accommodate the two configurations and have supplemented these results using solution studies and a molecular dynamics energetic analysis using a replica exchange methodology. Considering the previous examples of structural nonspecificity in coiled-coil peptides, the findings reported here not only emphasize the predisposition of the coiled-coil motif to adopt multiple configurations but also call attention to the associated risk that observed crytstal structures may not represent the only (or even the major) species present in solution.

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Figures

Figure 1

Figure 1

Crystal structures of the E20S variant (peptide 4) in the parallel and antiparallel tetrameric configurations. Side and top views show the parallel (a) and antiparallel (b) structures, highlighting core Leu (white) and Ile (blue) residues, with schematic diagrams showing interhelical packing interactions. c) Superposition of single helices from the antiparallel (blue) and parallel (beige) configurations. Backbone, β-carbons, and core residue heavy atoms are shown.

Figure 2

Figure 2

Comparison of hydrophobic packing interactions in the parallel and antiparallel E20S configurations. a) An overlay of the antiparallel (blue) and parallel (beige) E20S structures shows the shifted register for two helices in the antiparallel tetramer (with one helix omitted for clarity). Cα-Cβ bonds of hydrophobic core residues are shown, with Cβ atoms colored yellow, to illustrate the altered core geometry imposed by the antiparallel configuration. b) Cross-section of the same superposition showing the solvent-exposed surfaces of the antiparallel (blue mesh) and parallel (beige mesh) tetramers, along with side chain atoms for hydrophobic core residues. While the surfaces of the two assemblies are dissimilar, the core residues closely overlap.

Figure 3

Figure 3

Electrostatic interaction surfaces for the parallel and antiparallel coiled-coil tetramers. Dotted black lines are shown for potential polar intra- and interhelical interactions (< 4 Å) between surface residues for a) GCN4-pLI, b) E20S in the parallel configuration, c) E20S in the antiparallel configuration (K15 face), and d) E20S in the antiparallel configuration (S20 face). Positions 15 and 20 are colored yellow. Bridging water molecules are shown as red spheres.

Figure 4

Figure 4

Equilibrium disulfide exchange assay for E20S (

). a) Assuming the glycyl linkers allow random sorting of the terminal thiols, tetramers in the parallel configuration will form only parallel homodimeric disulfides, whereas antiparallel tetramers will also form the antiparallel heterodimeric disulfide. b) HPLC traces showing disulfide exchange over the course of the equilibration. The disulfide-bonded antiparallel heterodimer (Ns-sC, 30 μM) rearranges to form only the parallel homodimers (Ns-sN and Cs-sC). In an analogous experiment initiated with equimolar amounts of the parallel dimers, no change was observed after 88 h, indicating the system is at equilibrium. Peptide 12 has a larger extinction coefficient than peptide 11 because it contains an added tyrosine residue to facilitate HPLC separation. The small peak eluting between Ns-sN and Ns-sC is a disulfide adduct of glutathione to peptide 11.

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References

    1. Crick FHC. The packing of alpha-helices: simple coiled-coils. Acta Cryst. 1953;6:689–97.
    1. Alber T. How GCN4 binds DNA. Curr Biol. 1993;3:182–4. - PubMed
    1. O’Shea EK, Klemm JD, Kim PS, Alber T. X-ray structure of the GCN4 leucine zipper, a two-stranded, parallel coiled coil. Science. 1991;254:539–44. - PubMed
    1. Harbury PB, Zhang T, Kim PS, Alber T. A switch between two-, three, and four-stranded coiled coils in GCN4 leucine zipper mutants. Science. 1993;262:1401–7. - PubMed
    1. Harbury PB, Kim PS, Alber T. Crystal structure of an isoleucine-zipper trimer. Nature. 1994;371:80–3. - PubMed

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