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Quality control of disulfide bond formation in pilus subunits by the chaperone FimC

Nature Chemical Biology volume 8, pages 707–713 (2012)Cite this article

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Abstract

Type 1 pili from uropathogenic Escherichia coli are filamentous, noncovalent protein complexes mediating bacterial adhesion to the host tissue. All structural pilus subunits are homologous proteins sharing an invariant disulfide bridge. Here we show that disulfide bond formation in the unfolded subunits, catalyzed by the periplasmic oxidoreductase DsbA, is required for subunit recognition by the assembly chaperone FimC and for FimC-catalyzed subunit folding. FimC thus guarantees quantitative disulfide bond formation in each of the up to 3,000 subunits of the pilus. The X-ray structure of the complex between FimC and the main pilus subunit FimA and the kinetics of FimC-catalyzed FimA folding indicate that FimC accelerates folding of pilus subunits by lowering their topological complexity. The kinetic data, together with the measured in vivo concentrations of DsbA and FimC, predict an in vivo half-life of 2 s for oxidative folding of FimA in the periplasm.

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Figure 1: Schematic of type 1 pilus assembly by the chaperone-usher pathway.
Figure 2: Complex formation between FimAUox and FimC at pH 7.0 and 25 °C.
Figure 3: Stopped-flow fluorescence kinetics of FimAUred oxidation by DsbAox and subsequent FimAUox folding and complex formation with FimC (pH 7.0, 25 °C).
Figure 4: Kinetics of FimC-catalyzed folding of FimAUox at pH 7.0, 25 °C, showing that binding of FimAUox is the rate-limiting step in the formation of the native FimC-FimA complex.
Figure 5: Mechanism and in vivo kinetics of DsbA- and FimC-catalyzed folding of FimA.
Figure 6: 2.5-Å X-ray structure of the FimC–FimAt complex.

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Acknowledgements

This work was supported by the Swiss National Science Foundation (grants 310030B-138657 and 31003A-122095 to R.G.) and the Swiss Federal Institute of Technology Zürich within the framework of the Swiss National Center for Competence in Research Structural Biology Program. The PhD position of M.A.S. was supported by a grant from the Research Committee of the Paul Scherrer Institute (FK-05.08.1) to G.C.

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Author notes

  1. Oliv Eidam

    Present address: Present address: Department of Pharmaceutical Chemistry, University of California–San Francisco, San Francisco, California, USA.,

  2. Chasper Puorger and Martin A Schärer: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biology, Institute of Molecular Biology and Biophysics, Swiss Federal Institute of Technology Zurich, Zurich, Switzerland

    Maria D Crespo, Chasper Puorger & Rudi Glockshuber

  2. Laboratory of Biomolecular Research, Paul Scherrer Institute, Villigen, Switzerland

    Martin A Schärer & Guido Capitani

  3. Department of Biochemistry, University of Zurich, Zurich, Switzerland

    Oliv Eidam, Markus G Grütter & Guido Capitani

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  1. Maria D Crespo
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Contributions

The overall study was conceived and designed by M.D.C. and R.G. The biochemical experiments were performed by M.D.C. with contributions from C.P., M.A.S., O.E. and M.G.G., and G.C. contributed to the crystallographic structure determination, refinement and interpretation. M.D.C. and R.G. wrote the manuscript with the contribution of C.P., M.A.S. and G.C.

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Correspondence to Rudi Glockshuber.

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Crespo, M., Puorger, C., Schärer, M. et al. Quality control of disulfide bond formation in pilus subunits by the chaperone FimC. Nat Chem Biol 8, 707–713 (2012). https://doi.org/10.1038/nchembio.1019

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