Abstract

Proteins require an optimal balance of conformational flexibility and stability in their native environment to ensure their biological functions. A striking example is spidroins, spider silk proteins, which are stored at extremely high concentrations in soluble form, yet undergo amyloid-like aggregation during spinning. Here, we elucidate the stability of the highly soluble N-terminal domain (NT) of major ampullate spidroin 1 in the Escherichia coli cytosol as well as in inclusion bodies containing fibrillar aggregates. Surprisingly, we find that NT, despite being largely composed of amyloidogenic sequences, showed no signs of concentration-dependent aggregation. Using a novel intracellular hydrogen/deuterium exchange mass spectrometry (HDX-MS) approach, we reveal that NT adopts a tight fold in the E. coli cytosol and in this manner conceals its aggregation-prone regions by maintaining a tight fold under crowded conditions. Fusion of NT to the unstructured amyloid-forming A beta(40) peptide, on the other hand, results in the formation of fibrillar aggregates. However, HDX-MS indicates that the NT domain is only partially incorporated into these aggregates in vivo. We conclude that NT is able to control its aggregation to remain functional under the extreme conditions in the spider silk gland.

Keywords

hydrogen; deuterium exchange mass spectrometry; intracellular protein folding; protein aggregation; spider silk

Published in

FEBS Journal
2020, Volume: 287, number: 13, pages: 2823-2833
Publisher: WILEY

SLU Authors

• Rising, Anna

  • Department of Animal Biosciences, Swedish University of Agricultural Sciences
    
  • Karolinska Institute

UKÄ Subject classification

Biochemistry and Molecular Biology


Publication identifier

DOI: https://doi.org/10.1111/febs.15169

Permanent link to this page (URI)

https://res.slu.se/id/publ/103365