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Three-dimensional map of the plasma membrane H+-ATPase in the open conformation

Abstract

The H+-ATPase from the plasma membrane of Neurospora crassa is an integral membrane protein of relative molecular mass 100K, which belongs to the P-type ATPase family that includes the plasma membrane Na+/K+-ATPase and the sarcoplasmic reticulum Ca2+-ATPase. The H+-ATPase pumps protons across the cell's plasma membrane using ATP as an energy source, generating a membrane potential in excess of 200 mV (13). Despite the importance of P-type ATPases in controlling membrane potential and intracellular ion concentrations, little is known about the molecular mechanism they use for ion transport. This is largely due to the difficulty in growing well ordered crystals and the resulting lack of detail in the three-dimensional structure of these large membrane proteins. We have now obtained a three-dimensional map of the H+-ATPase by electron crystallography of two-dimensional crystals grown directly on electron microscope grids. At an in-plane resolution of 8 Å, this map reveals ten membrane-spanning α-helices in the membrane domain, and four major cytoplasmic domains in the open conformation of the enzyme without bound ligands.

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Figure 1: Phases (top panels) and amplitudes of two lattice lines.
Figure 2: Three-dimensional map of the H+-ATPase.
Figure 3: Side view of the H+-ATPase monomer, showing the average of two monomers related by non-crystallographic two-fold symmetry.
Figure 4: The transmembrane region.

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Acknowledgements

We thank M. Radermacher for help with the program SPIDER, D. Mills and J. Vonck for help with the JEOL 3000 SFF, and J. Postma for assistance with computer graphics. W.K. gratefully acknowledges financial support from the Deutsche Forschungsgemeinschaft. G.A.S. was supported by a grant from USPHS NIH.

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Correspondence to Werner Kühlbrandt.

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Auer, M., Scarborough, G. & Kühlbrandt, W. Three-dimensional map of the plasma membrane H+-ATPase in the open conformation. Nature 392, 840–843 (1998). https://doi.org/10.1038/33967

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