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Self-assembly of spider silk proteins is controlled by a pH-sensitive relay

Nature volume 465, pages 236–238 (2010)Cite this article

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Abstract

Nature’s high-performance polymer, spider silk, consists of specific proteins, spidroins, with repetitive segments flanked by conserved non-repetitive domains1,2. Spidroins are stored as a highly concentrated fluid dope. On silk formation, intermolecular interactions between repeat regions are established that provide strength and elasticity3,4. How spiders manage to avoid premature spidroin aggregation before self-assembly is not yet established. A pH drop to 6.3 along the spider’s spinning apparatus, altered salt composition and shear forces are believed to trigger the conversion to solid silk, but no molecular details are known. Miniature spidroins consisting of a few repetitive spidroin segments capped by the carboxy-terminal domain form metre-long silk-like fibres irrespective of pH5. We discovered that incorporation of the amino-terminal domain of major ampullate spidroin 1 from the dragline of the nursery web spider Euprosthenops australis (NT) into mini-spidroins enables immediate, charge-dependent self-assembly at pH values around 6.3, but delays aggregation above pH 7. The X-ray structure of NT, determined to 1.7 Å resolution, shows a homodimer of dipolar, antiparallel five-helix bundle subunits that lack homologues. The overall dimeric structure and observed charge distribution of NT is expected to be conserved through spider evolution and in all types of spidroins. Our results indicate a relay-like mechanism through which the N-terminal domain regulates spidroin assembly by inhibiting precocious aggregation during storage, and accelerating and directing self-assembly as the pH is lowered along the spider’s silk extrusion duct.

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Figure 1: pH-dependent assembly of NT and mini-spidroins.
Figure 2: Structural and conserved features of spidroin N-terminal domain.

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Acknowledgements

This work was supported by grants (to J.J., M.H. and S.D.K.) from the Swedish Research Council and FORMAS. We thank E. Andersson, K. Tars and D. Ericsson for collecting the high resolution wild-type X-ray diffraction data set, H. Bramfeldt and C. Aulin for help with collecting scanning electron microscopy images, L. Serpell, I. Andersson, A. Zavialov and T. Härd for comments on the manuscript, and a number of other colleagues for discussions. We thank the ESRF (Grenoble, France) beamline staff for help during data collection.

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Author notes

  1. Glareh Askarieh and My Hedhammar: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Chemistry, Oslo University, 1033 Blindern, 0315 Oslo, Norway,

    Glareh Askarieh

  2. Department of Molecular Biology, Uppsala BioCenter, SLU, Biomedical Centre, P.O. Box 590, SE-751 24 Uppsala, Sweden,

    Glareh Askarieh & Stefan D. Knight

  3. Department of Anatomy, Physiology and Biochemistry, SLU, Biomedical Centre, P.O. Box 575, SE-751 23 Uppsala, Sweden,

    My Hedhammar, Kerstin Nordling, Anna Rising & Jan Johansson

  4. Department of Biochemistry and Molecular Biology I & CIBER Enfermedades Respiratorias, Complutense University of Madrid, 28040-Madrid, Spain

    Alejandra Saenz & Cristina Casals

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  1. Glareh Askarieh
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Contributions

G.A. and M.H. contributed equally to this work. G.A., M.H. performed experiments and wrote the paper, K.N., A.S. performed experiments, C.C., A.R., J.J., S.D.K. discussed experiments and wrote the paper.

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Correspondence to Jan Johansson or Stefan D. Knight.

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Competing interests

M.H., K.N., A.R. and J.J. own stocks and are funded by Spiber Technologies AB, a company that aims to commercialize recombinant spider silk for biomedical applications.

Additional information

X-ray crystallographic coordinates and structure factors have been deposited in the RCSB Protein Data Bank (PDB) with PDB ID codes 3LR2, 3LR6, 3LR8, 3LRD.

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This file contains Supplementary Figures 1-8 with legends and Supplementary Tables 1-2. (PDF 607 kb)

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Askarieh, G., Hedhammar, M., Nordling, K. et al. Self-assembly of spider silk proteins is controlled by a pH-sensitive relay. Nature 465, 236–238 (2010). https://doi.org/10.1038/nature08962

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