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Structural analysis of a prototypical ATPase from the type III secretion system

Nature Structural & Molecular Biology volume 14, pages 131–137 (2007)Cite this article

Abstract

The type III secretion system (T3SS) ATPase is the conserved and essential inner-membrane component involved in the initial stages of selective secretion of specialized T3SS virulence effector proteins from the bacterial cytoplasm through to the infected host cell, a process crucial to subsequent pathogenicity. Here we present the 1.8-Å-resolution crystal structure of the catalytic domain of the prototypical T3SS ATPase EscN from enteropathogenic Escherichia coli (EPEC). Along with in vitro and in vivo mutational analysis, our data show that the T3SS ATPases share similarity with the F1 ATPases but have important structural and sequence differences that dictate their unique secretory role. We also show that T3SS ATPase activity is dependent on EscN oligomerization and describe the molecular features and possible functional implications of a hexameric ring model.

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Figure 1: The overall architecture of EscNΔ102-V393P.
Figure 2: Biochemical data for the V393P mutant.
Figure 3: The ATP-binding pocket in EscN, as delineated by soaking of the nonhydrolyzable ATP analog AMP-PNP.
Figure 4: Biochemical analysis of the R366D mutant.
Figure 5: Functional mutants of EscN.
Figure 6: The hexameric T3SS and F1 ATPases.

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Acknowledgements

We thank A.L. Lovering, C.K. Yip and P.I. Lario for discussions, C.K. Yip for performing the static light-scattering analysis, N.A. Thomas for reagents and the staff at Advanced Light Source beamline 8.2.2 for data collection time and assistance. R.Z. is supported by postdoctoral fellowships from Izaak Walton Killam Research, the Michael Smith Foundation for Health Research (MSFHR) and the Canadian Institutes of Health Research (CIHR). N.C.J.S. and B.B.F. thank the Howard Hughes International Scholar program and the CIHR for funding. N.C.J.S. also thanks the MSFHR and the Canada Foundation for Innovation for infrastructure funding support. N.C.J.S. is an MSFHR Senior Scholar and CIHR Investigator.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology and the Center for Blood Research, University of British Columbia, 2350 Health Sciences Mall, Vancouver, V6T 1Z3, British Columbia, Canada

    Raz Zarivach, Marija Vuckovic & Natalie C J Strynadka

  2. Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, V6T 1Z4, British Columbia, Canada

    Wanyin Deng & B Brett Finlay

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  1. Raz Zarivach
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Correspondence to Natalie C J Strynadka.

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Supplementary information

Supplementary Fig. 1

EscN structure. (PDF 1305 kb)

Supplementary Fig. 2

Multiple sequence alignment. (PDF 1766 kb)

Supplementary Table 1

Structural comparison table. (PDF 2042 kb)

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Zarivach, R., Vuckovic, M., Deng, W. et al. Structural analysis of a prototypical ATPase from the type III secretion system. Nat Struct Mol Biol 14, 131–137 (2007). https://doi.org/10.1038/nsmb1196

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