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Crystal structure of a rhomboid family intramembrane protease

Abstract

Escherichia coli GlpG is an integral membrane protein that belongs to the widespread rhomboid protease family. Rhomboid proteases, like site-2 protease (S2P) and γ-secretase, are unique in that they cleave the transmembrane domain of other membrane proteins. Here we describe the 2.1 Å resolution crystal structure of the GlpG core domain. This structure contains six transmembrane segments. Residues previously shown to be involved in catalysis, including a Ser–His dyad, and several water molecules are found at the protein interior at a depth below the membrane surface. This putative active site is accessible by substrate through a large ‘V-shaped’ opening that faces laterally towards the lipid, but is blocked by a half-submerged loop structure. These observations indicate that, in intramembrane proteolysis, the scission of peptide bonds takes place within the hydrophobic environment of the membrane bilayer. The crystal structure also suggests a gating mechanism for GlpG that controls substrate access to its hydrophilic active site.

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Figure 1: The membrane topology of a rhomboid protease and its substrate.
Figure 2: The overall structure of GlpG.
Figure 3: The membrane-embedded active site of rhomboid protease.
Figure 4: The structure of the lateral gate.
Figure 5: A possible mechanism of rhomboid-catalysed intramembrane proteolysis.

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Acknowledgements

We thank V. Stojanoff, H. Robinson, A. Saxena and A. Héroux at BNL NSLS beamlines for help; T. Boggon and J. Schlessinger for sharing the crystallization robot in their laboratories; and B. Turk for sharing the fluorescence spectrometer in his laboratory. X-ray diffraction data for this study were measured at beamlines X6A, X29 and X26C of NSLS. Financial support comes principally from the US Department of Energy, and from the National Institutes of Health. This work was supported by a New Scholar Award in Aging from the Ellison Medical Foundation (to Y.H.) and a gift from the Neuroscience Education and Research Foundation (to Y.H.). Author Contributions Y.W. and Y.H. purified and characterized GlpG in various detergents. Y.W. crystallized GlpG. Y.H. and Y.W. solved the structure of GlpG and wrote the paper. Y.Z. screened the expression of many constructs and conducted some initial biochemical characterizations.

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Correspondence to Ya Ha.

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The atomic coordinates of GlpG have been deposited in the Protein Data Bank (accession number 2IC8), and will be released upon publication of the paper. Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests.

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Wang, Y., Zhang, Y. & Ha, Y. Crystal structure of a rhomboid family intramembrane protease. Nature 444, 179–180 (2006). https://doi.org/10.1038/nature05255

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