High Intracellular Stability of the Spidroin N-terminal Domain in Spite of Abundant Amyloidogenic Segments Revealed by In-Cell Hydrogen/Deuterium Exchange Mass Spectrometry

High Intracellular Stability of the Spidroin N-terminal Domain in Spite of Abundant Amyloidogenic Segments Revealed by In-Cell Hydrogen/Deuterium Exchange Mass Spectrometry

The FEBS Journal

10.1111/febs.15169

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

Margit Kaldmäe, Axel Leppert, Gefei Chen, Medoune Sarr, Cagla Sahin, Kerstin Nordling, Nina Kronqvist, Marta Gonzalvo-Ulla, Nicolas Fritz, Axel Abelein, Sonia Laίn, Henrik Biverstal, Hans Jornvall, David P. Lane, Anna Rising, Jan Johansson, Michael Landreh

hydrogen/deuterium exchange mass spectrometry, intracellular protein folding, protein aggregation, spider silk.

01 December 2019

Kaldmäe, M., Leppert, A., Chen, G., Sarr, M., Sahin, C., Nordling, K., Kronqvist, N., Gonzalvo‐Ulla, M., Fritz, N., Abelein, A., Laίn, S., Biverstål, H., Jörnvall, H., Lane, D.P., Rising, A., Johansson, J. and Landreh, M. (2020), High intracellular stability of the spidroin N‐terminal domain in spite of abundant amyloidogenic segments revealed by in‐cell hydrogen/deuterium exchange mass spectrometry. FEBS J. doi:10.1111/febs.15169

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 Aβ40 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.

http://creativecommons.org/licenses/by-nc/4.0/

ML is supported by an Ingvar Carlsson Award from the Swedish Foundation for Strategic Research, a KI Faculty-funded Career Position, a KI-StratNeuro Starting Grant and a VR Starting Grant (2019-01961). CS is supported by a Novo Nordisk Foundation Postdoctoral Fellowship (NNF19OC0055700). SL is supported by Barncancerfonden grant TJ2014-0013. DPL is supported by VR grant 2013_08807. JJ and AR acknowledge support from the Center for Innovative Medicine at Karolinska Institutet (CIMED), VR and Vinnova. ML gratefully acknowledges technical support from MS Vision, NL.

https://oar.a-star.edu.sg/communities-collections/articles/15990

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