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A de novo peptide hexamer with a mutable channel

Abstract

The design of new proteins that expand the repertoire of natural protein structures represents a formidable challenge. Success in this area would increase understanding of protein structure and present new scaffolds that could be exploited in biotechnology and synthetic biology. Here we describe the design, characterization and X-ray crystal structure of a new coiled-coil protein. The de novo sequence forms a stand-alone, parallel, six-helix bundle with a channel running through it. Although lined exclusively by hydrophobic leucine and isoleucine side chains, the 6-Å channel is permeable to water. One layer of leucine residues within the channel is mutable, accepting polar aspartic acid and histidine side chains, which leads to subdivision and organization of solvent within the lumen. Moreover, these mutants can be combined to form a stable and unique (Asp-His)3 heterohexamer. These new structures provide a basis for engineering de novo proteins with new functions.

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Figure 1: A coiled-coil helical wheel and parallel coiled-coil assemblies.
Figure 2: Helicity and oligomerization of CC-Tet and CC-Hex.
Figure 3: Structure of a parallel coiled-coil hexamer.
Figure 4: KIH interactions in the CC-Hex structure.
Figure 5: X-ray crystal structures of mutant CC-Hex channels.

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Acknowledgements

We thank members of the Woolfson group, in particular C. Armstrong and J. Fletcher, for discussions, and H. Bayley, A. Lupas and J. Spencer for comments on the manuscript. We are grateful to the Biotechnology and Biological Sciences Research Council of the UK for grants to D.N.W. and P.J.B. (BB/G008833/1) and to R.L.B. and D.N.W. (BB/F007256/1).

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Contributions

A.R.T., B.C., G.J.B. and D.N.W. designed the peptide sequences; B.C. and A.R.T. synthesized the peptides; B.C., A.L.B. and A.R.T. performed the solution-phase biophysics; N.R.Z., B.C. and R.L.B. solved the peptide X-ray crystal structures; G.J.B. performed the bioinformatics; M.B., A.R.T. and D.N.W. conducted the SOCKET and TWISTER analyses; R.B.S., N.L. and D.N.W. conceived and conducted the computational modeling; P.J.B., R.L.B. and D.N.W. conceived and supervised the experimental program; D.N.W. wrote the manuscript.

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Correspondence to R Leo Brady or Derek N Woolfson.

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Zaccai, N., Chi, B., Thomson, A. et al. A de novo peptide hexamer with a mutable channel. Nat Chem Biol 7, 935–941 (2011). https://doi.org/10.1038/nchembio.692

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