Designed peptides that assemble into cross-α amyloid-like structures

Abstract

Amyloids adopt ‘cross-β’ structures composed of long, twisted fibrils with β-strands running perpendicular to the fibril axis. Recently, a toxic peptide was proposed to form amyloid-like cross-α structures in solution, with a planar bilayer-like assembly observed in the crystal structure. Here we crystallographically characterize designed peptides that assemble into spiraling cross-α amyloid-like structures, which resemble twisted β-amyloid fibrils. The peptides form helical dimers, stabilized by packing of small and apolar residues, and the dimers further assemble into cross-α amyloid-like fibrils with superhelical pitches ranging from 170 Å to 200 Å. When a small residue that appeared critical for packing was converted to leucine, it resulted in structural rearrangement to a helical polymer. Fluorescently tagged versions of the designed peptides form puncta in mammalian cells, which recover from photobleaching with markedly different kinetics. These structural folds could be potentially useful for directing in vivo protein assemblies with predetermined spacing and stabilities.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1: The amyloid-like structure of αAmmem.
Fig. 2: Design of cross-α amyloid-like assembly.
Fig. 3: The aggregation behavior and fibril formation of the water-soluble peptides and the crystal structures of the cross-α amyloid-like fibrils.
Fig. 4: Packing of αAmL as a helical polymer composed of helix tetramers and its configurational relationship with the cross-α amyloid-like assembly of αAmG and αAmS.
Fig. 5: EGFP-tagged αAm peptides form inclusions in the cytosol of mammalian cells; αAmG, αAmA and αAmS form a solid-like phase, whereas αAmL is more mobile.

References

  1. 1.

    Gazit, E. Self-assembled peptide nanostructures: the design of molecular building blocks and their technological utilization. Chem. Soc. Rev. 36, 1263–1269 (2007).

    Article  CAS  Google Scholar 

  2. 2.

    Shigemitsu, H. & Hamachi, I. Design strategies of stimuli-responsive supramolecular hydrogels relying on structural analyses and cell-mimicking approaches. Acc. Chem. Res. 50, 740–750 (2017).

    Article  CAS  Google Scholar 

  3. 3.

    Shu, J. Y., Panganiban, B. & Xu, T. Peptide-polymer conjugates: from fundamental science to application. Annu. Rev. Phys. Chem. 64, 631–657 (2013).

    Article  CAS  Google Scholar 

  4. 4.

    Riek, R. & Eisenberg, D. S. The activities of amyloids from a structural perspective. Nature 539, 227–235 (2016).

    Article  Google Scholar 

  5. 5.

    Prusiner, S. B. Cell biology. A unifying role for prions in neurodegenerative diseases. Science 336, 1511–1513 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Sacchettini, J. C. & Kelly, J. W. Therapeutic strategies for human amyloid diseases. Nat. Rev. Drug Discov. 1, 267–275 (2002).

    Article  CAS  Google Scholar 

  7. 7.

    Toyama, B. H. & Weissman, J. S. Amyloid structure: conformational diversity and consequences. Annu. Rev. Biochem. 80, 557–585 (2011).

    Article  CAS  Google Scholar 

  8. 8.

    Knowles, T. P. & Buehler, M. J. Nanomechanics of functional and pathological amyloid materials. Nat. Nanotechnol. 6, 469–479 (2011).

    Article  CAS  Google Scholar 

  9. 9.

    Rufo, C. M. et al. Short peptides self-assemble to produce catalytic amyloids. Nat. Chem. 6, 303–309 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Makhlynets, O. V., Gosavi, P. M. & Korendovych, I. V. Short self-assembling peptides are able to bind to copper and activate oxygen. Angew. Chem. Int. Edn Engl. 55, 9017–9020 (2016).

    Article  CAS  Google Scholar 

  11. 11.

    Tena-Solsona, M. et al. Emergent catalytic behavior of self-assembled low molecular weight peptide-based aggregates and hydrogels. Chemistry 22, 6687–6694 (2016).

    Article  CAS  Google Scholar 

  12. 12.

    Friedmann, M. P. et al. Towards prebiotic catalytic amyloids using high throughput screening. PLoS One 10, e0143948 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Childers, W. S., Ni, R., Mehta, A. K. & Lynn, D. G. Peptide membranes in chemical evolution. Curr. Opin. Chem. Biol. 13, 652–659 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Tayeb-Fligelman, E. et al. The cytotoxic Staphylococcus aureus PSMα3 reveals a cross-α amyloid-like fibril. Science 355, 831–833 (2017).

    Article  Google Scholar 

  15. 15.

    Privé, G. G., Anderson, D. H., Wesson, L., Cascio, D. & Eisenberg, D. Packed protein bilayers in the 0.90 A resolution structure of a designed alpha helical bundle. Protein Sci. 8, 1400–1409 (1999).

    Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Patterson, W. R., Anderson, D. H., DeGrado, W. F., Cascio, D. & Eisenberg, D. Centrosymmetric bilayers in the 0.75 A resolution structure of a designed alpha-helical peptide, d,l-Alpha-1. Protein Sci. 8, 1410–1422 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. 17.

    Egelman, E. H. et al. Structural plasticity of helical nanotubes based on coiled-coil assemblies. Structure 23, 280–289 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Brunette, T. J. et al. Exploring the repeat protein universe through computational protein design. Nature 528, 580–584 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Main, E. R., Jackson, S. E. & Regan, L. The folding and design of repeat proteins: reaching a consensus. Curr. Opin. Struct. Biol. 13, 482–489 (2003).

    Article  CAS  Google Scholar 

  20. 20.

    Plückthun, A. Designed ankyrin repeat proteins (DARPins): binding proteins for research, diagnostics, and therapy. Annu. Rev. Pharmacol. Toxicol. 55, 489–511 (2015).

    Article  CAS  Google Scholar 

  21. 21.

    Joh, N. H. et al. De novo design of a transmembrane Zn2+-transporting four-helix bundle. Science 346, 1520–1524 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Zhang, S.-Q. et al. De novo design of tetranuclear transition metal clusters stabilized by hydrogen-bonded networks in helical bundles. J. Am. Chem. Soc. 140, 1294–1304 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. 23.

    Eisenberg, D. S. & Sawaya, M. R. Structural studies of amyloid proteins at the molecular level. Annu. Rev. Biochem. 86, 69–95 (2017).

    Article  CAS  Google Scholar 

  24. 24.

    Szczepaniak, K., Lach, G., Bujnicki, J. M. & Dunin-Horkawicz, S. Designability landscape reveals sequence features that define axial helix rotation in four-helical homo-oligomeric antiparallel coiled-coil structures. J. Struct. Biol. 188, 123–133 (2014).

    Article  CAS  Google Scholar 

  25. 25.

    Banner, D. W., Kokkinidis, M. & Tsernoglou, D. Structure of the ColE1 rop protein at 1.7 A resolution. J. Mol. Biol. 196, 657–675 (1987).

    Article  CAS  Google Scholar 

  26. 26.

    Sanders, D. W. et al. Distinct tau prion strains propagate in cells and mice and define different tauopathies. Neuron 82, 1271–1288 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. 27.

    Prusiner, S. B. et al. Evidence for α-synuclein prions causing multiple system atrophy in humans with parkinsonism. Proc. Natl. Acad. Sci. USA 112, E5308–E5317 (2015).

    Article  CAS  Google Scholar 

  28. 28.

    Thompson, K. E., Bashor, C. J., Lim, W. A. & Keating, A. E. SYNZIP protein interaction toolbox: in vitro and in vivo specifications of heterospecific coiled-coil interaction domains. ACS Synth. Biol. 1, 118–129 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Fletcher, J. M. et al. A basis set of de novo coiled-coil peptide oligomers for rational protein design and synthetic biology. ACS Synth. Biol. 1, 240–250 (2012).

    Article  CAS  Google Scholar 

  30. 30.

    Banani, S. F., Lee, H. O., Hyman, A. A. & Rosen, M. K. Biomolecular condensates: organizers of cellular biochemistry. Nat. Rev. Mol. Cell Biol. 18, 285–298 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Kim, C. A., Sawaya, M. R., Cascio, D., Kim, W. & Bowie, J. U. Structural organization of a Sex-comb-on-midleg/polyhomeotic copolymer. J. Biol. Chem. 280, 27769–27775 (2005).

    Article  CAS  Google Scholar 

  32. 32.

    Kim, C. A. & Bowie, J. U. SAM domains: uniform structure, diversity of function. Trends Biochem. Sci. 28, 625–628 (2003).

    Article  CAS  Google Scholar 

  33. 33.

    Wu, H. & Fuxreiter, M. The structure and dynamics of higher-order assemblies: amyloids, signalosomes, and granules. Cell 165, 1055–1066 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Kuhlman, B. & Baker, D. Native protein sequences are close to optimal for their structures. Proc. Natl. Acad. Sci. USA 97, 10383–10388 (2000).

    Article  CAS  Google Scholar 

  35. 35.

    Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  Google Scholar 

  36. 36.

    Kabsch, W. Integration, scaling, space-group assignment and post-refinement. Acta Crystallogr. D Biol. Crystallogr. 66, 133–144 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. 38.

    McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Murshudov, G. N. et al. REFMAC5 for the refinement of macromolecular crystal structures. Acta Crystallogr. D Biol. Crystallogr. 67, 355–367 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Winn, M. D. et al. Overview of the CCP4 suite and current developments. Acta Crystallogr. D Biol. Crystallogr. 67, 235–242 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structuresolution. Acta Crystallogr. D Biol. Crystallogr. 66, 213–221 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank D. Bulkley, P. Jin, S. Li, X. Liu, N. Polizzi, N. Schmidt and H. Wu for technical help. This work was primarily supported by NIH grant R35GM122603 to W.F.D., with additional support from the NSF (CHE1413295) for the MRSEC program to the LRSM at the University of Pennsylvania. H.T.K. was supported by a Ruth L. Kirschstein NRSA Postdoctoral Fellowship (F32GM125217). Y.L. was supported by a Howard Hughes Medical Institute-Helen Hay Whitney Foundation Postdoctoral Fellowship.

Author information

Affiliations

Authors

Contributions

S.-Q.Z. and W.F.D. conceived the project. S.-Q.Z. designed all the peptide sequences and performed in vitro experiments with H.T.K., M.L. and Y.L. H.H. and J.Y. conducted cellular in vivo experiments. L.L. solved and refined all the crystal structures. S.-Q.Z., H.H, J.Y., H.T.K., Y.L., X.S., L.L. and W.F.D. analyzed the data. S.-Q.Z. and W.F.D. prepared the manuscript with contributions from all the authors.

Corresponding authors

Correspondence to Xiaokun Shu or Lijun Liu or William F. DeGrado.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Text and Figures

Supplementary Table 1–2, Supplementary Figures 1–11

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Zhang, S., Huang, H., Yang, J. et al. Designed peptides that assemble into cross-α amyloid-like structures. Nat Chem Biol 14, 870–875 (2018). https://doi.org/10.1038/s41589-018-0105-5

Download citation

Further reading