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Amyloid-like fibrils of ribonuclease A with three-dimensional domain-swapped and native-like structure

Nature volume 437, pages 266–269 (2005)Cite this article

Abstract

Amyloid or amyloid-like fibrils are elongated, insoluble protein aggregates, formed in vivo1 in association with neurodegenerative diseases or in vitro2 from soluble native proteins, respectively. The underlying structure of the fibrillar or ‘cross-β’ state has presented long-standing, fundamental puzzles of protein structure. These include whether fibril-forming proteins have two structurally distinct stable states, native and fibrillar, and whether all or only part of the native protein refolds as it converts to the fibrillar state. Here we show that a designed amyloid-like fibril of the well-characterized enzyme RNase A contains native-like molecules capable of enzymatic activity. In addition, these functional molecular units are formed from a core RNase A domain and a swapped complementary domain. These findings are consistent with the zipper-spine model3 in which a cross-β spine is decorated with three-dimensional domain-swapped functional units, retaining native-like structure.

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Figure 1: RNase A monomer and C-terminal domain-swapped dimer and the 3D domain-swapped zipper-spine model.
Figure 2: Properties of the RNase A amyloid-like fibrils.
Figure 3: Fluorescence-based activity assay to examine whether the RNase A amyloid-like fibrils are domain-swapped.
Figure 4: Domain-swapped zipper-spine model for the RNase A protofibril.

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Acknowledgements

We thank M. Phillips for electron microscopy, M. Bennett, T. Yeates and D. H. Anderson for discussion, and M. Nebohacova and L. Simpson for assistance with blue-native gels. This study received financial support from the NIH, NSF and the Howard Hughes Medical Institute.

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Author notes

  1. Yanshun Liu

    Present address: Division of Chemistry and Chemical Engineering, Caltech, Pasadena, California, 91125, USA

Authors and Affiliations

  1. Howard Hughes Medical Institute, UCLA-DOE Institute for Genomics and Proteomics, UCLA, Box 951570, California, 90095-1570, Los Angeles, USA

    Shilpa Sambashivan, Yanshun Liu, Michael R. Sawaya, Mari Gingery & David Eisenberg

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  1. Shilpa Sambashivan
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Corresponding author

Correspondence to David Eisenberg.

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Competing interests

The structure of Q10-H119A RNase A has been deposited in the Protein Data Bank with accession code 2APQ. The model of Fig. 4 has been deposited in the Protein Data Bank with accession code 2APU. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Notes

This file includes Supplementary Methods, additional control experiments and legends for Supplementary Table S1, Supplementary Figure S1 and Supplementary Figure S2. (DOC 1824 kb)

Supplementary Table S2

Data Collection for Q10-H119A (DOC 28 kb)

Supplementary Figure S1

Cross- diffraction from partially oriented Q10-H119A RNase A fibrils. (JPG 62 kb)

Supplementary Figure S2

Silver-stained, non-denaturing gel comparing lyophilized Q10 -expanded RNase A with lyophilized wild-type RNase A. (JPG 79 kb)

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Sambashivan, S., Liu, Y., Sawaya, M. et al. Amyloid-like fibrils of ribonuclease A with three-dimensional domain-swapped and native-like structure. Nature 437, 266–269 (2005). https://doi.org/10.1038/nature03916

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