Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment revealed by Cryo-EM and hydrogen/deuterium exchange mass spectrometry

Ross, J.; McIver, Z.; Lambert, T.; Piergentili, C.; Gallagher, K. J.; Bird, J. E.; Cruickshank, F. L.; Zarazua-Arvizu, E.; Horsfall, L. E.; Waldron, K. J.; Wilson, M. D.; Mackay, C. L.; Basle, A.; Clarke, D. J.; Marles-Wright, J.

doi:10.1101/2021.04.15.439977

bioRxiv · 10.1101/2021.04.15.439977

Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment revealed by Cryo-EM and hydrogen/deuterium exchange mass spectrometry

Ross, J.·McIver, Z.·Lambert, T.·Piergentili, C.·Gallagher, K. J.·Bird, J. E.·Cruickshank, F. L.·Zarazua-Arvizu, E.·Horsfall, L. E.·Waldron, K. J.·Wilson, M. D.·Mackay, C. L.·Basle, A.·Clarke, D. J.·Marles-Wright, J.

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Abstract

Encapsulins are protein nanocompartments that house various cargo enzymes, including a family of decameric ferritin-like proteins. Here, we study a recombinant Haliangium ochraceum encapsulin:encapsulated ferritin complex using electron cryo-microscopy and hydrogen/deuterium exchange mass spectrometry to gain insight into the structural relationship between the encapsulin shell and its protein cargo. An asymmetric single particle reconstruction reveals four encapsulated ferritin decamers in a tetrahedral arrangement within the encapsulin nanocompartment. This leads to a symmetry mismatch between the protein cargo and the icosahedral encapsulin shell. The encapsulated ferritin decamers are offset from the interior face of the encapsulin shell. Using HDX-MS, we observed dynamic behavior of the major five-fold pore in the encapsulin shell and show the pore opening via the movement of the encapsulin A-domain. These data will accelerate efforts to engineer the encapsulation of heterologous cargo proteins and to alter the permeability of the encapsulin shell via pore modifications. TeaserCryo-EM and HDX-MS analysis of an encapsulin nanocompartment shows that the pores at the five-fold icosahedral vertex of the shell are flexible.

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Ross, J., McIver, Z., Lambert, T., Piergentili, C., Gallagher, K. J., Bird, J. E., Cruickshank, F. L., Zarazua-Arvizu, E., Horsfall, L. E., Waldron, K. J., Wilson, M. D., Mackay, C. L., Basle, A., Clarke, D. J., Marles-Wright, J.. 2021-04-15. Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment revealed by Cryo-EM and hydrogen/deuterium exchange mass spectrometry. https://doi.org/10.1101/2021.04.15.439977

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