5VOX

Yeast V-ATPase in complex with Legionella pneumophila effector SidK (rotational state 1)

Summary for 5VOX

• Entry DOI: 10.2210/pdb5vox/pdb
• Related: 5VOY 5VOZ
• EMDB information: 8724 8725 8726
• Descriptor: V-type proton ATPase catalytic subunit A,V-type proton ATPase catalytic subunit A, V-type proton ATPase subunit c'', V-type proton ATPase subunit c', ... (16 entities in total)
• Functional Keywords: v-atpase, sidk, rotational state 1, hydrolase
• Biological source: Legionella pneumophila subsp. pneumophila ATCC 43290; More
• Total number of polymer chains: 33
• Total formula weight: 1151450.73

Authors

Zhao, J. (deposition date: 2017-05-03, release date: 2017-06-21, Last modification date: 2024-03-13)

Primary citation

Zhao, J.,Beyrakhova, K.,Liu, Y.,Alvarez, C.P.,Bueler, S.A.,Xu, L.,Xu, C.,Boniecki, M.T.,Kanelis, V.,Luo, Z.Q.,Cygler, M.,Rubinstein, J.L.
Molecular basis for the binding and modulation of V-ATPase by a bacterial effector protein.
PLoS Pathog., 13:e1006394-e1006394, 2017

PubMed Abstract: Intracellular pathogenic bacteria evade the immune response by replicating within host cells. Legionella pneumophila, the causative agent of Legionnaires' Disease, makes use of numerous effector proteins to construct a niche supportive of its replication within phagocytic cells. The L. pneumophila effector SidK was identified in a screen for proteins that reduce the activity of the proton pumping vacuolar-type ATPases (V-ATPases) when expressed in the yeast Saccharomyces cerevisae. SidK is secreted by L. pneumophila in the early stages of infection and by binding to and inhibiting the V-ATPase, SidK reduces phagosomal acidification and promotes survival of the bacterium inside macrophages. We determined crystal structures of the N-terminal region of SidK at 2.3 Å resolution and used single particle electron cryomicroscopy (cryo-EM) to determine structures of V-ATPase:SidK complexes at ~6.8 Å resolution. SidK is a flexible and elongated protein composed of an α-helical region that interacts with subunit A of the V-ATPase and a second region of unknown function that is flexibly-tethered to the first. SidK binds V-ATPase strongly by interacting via two α-helical bundles at its N terminus with subunit A. In vitro activity assays show that SidK does not inhibit the V-ATPase completely, but reduces its activity by ~40%, consistent with the partial V-ATPase deficiency phenotype its expression causes in yeast. The cryo-EM analysis shows that SidK reduces the flexibility of the A-subunit that is in the 'open' conformation. Fluorescence experiments indicate that SidK binding decreases the affinity of V-ATPase for a fluorescent analogue of ATP. Together, these results reveal the structural basis for the fine-tuning of V-ATPase activity by SidK.

PubMed: 28570695
DOI: 10.1371/journal.ppat.1006394

Experimental method

ELECTRON MICROSCOPY (6.8 Å)