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Prolyl isomerization as a molecular timer in phage infection

Nature Structural & Molecular Biology volume 12, pages 619–623 (2005)Cite this article

Abstract

Prolyl cis-trans isomerizations are intrinsically slow reactions and known to be rate-limiting in many protein folding reactions. Here we report that a proline is used as a molecular timer in the infection of Escherichia coli cells by the filamentous phage fd. The phage is activated for infection by the disassembly of the two N-terminal domains, N1 and N2, of its gene-3-protein, which is located at the phage tip. Pro213, in the hinge between N1 and N2, sets a timer for the infective state. The timer is switched on by cis-to-trans and switched off by the unusually slow trans-to-cis isomerization of the Gln212-Pro213 peptide bond. The switching rate and thus the infectivity of the phage are determined by the local sequence around Pro213, and can be tuned by mutagenesis.

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Figure 1: Function of G3P in phage infection.
Figure 2: Competitions between different phage variants for the infection of E. coli.
Figure 3: Loss of phage infectivity follows proline-limited domain closing.
Figure 4: The slow isomerization at Pro213 is determined by the local sequence.

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References

  1. Grathwohl, C. & Wüthrich, K. NMR studies of the rates of proline cis-trans isomerization in oligopeptides. Biopolymers 20, 2623–2633 (1981).

    Article  CAS  Google Scholar 

  2. Stein, R.L. Mechanism of enzymatic and nonenzymatic prolyl cis-trans isomerization. Adv. Protein Chem. 44, 1–24 (1993).

    Article  CAS  Google Scholar 

  3. Fischer, G. Chemical aspects of peptide bond isomerisation. Chem. Soc. Rev. 29, 119–127 (2000).

    Article  CAS  Google Scholar 

  4. Reimer, U. et al. Side-chain effects on peptidyl-prolyl cis/trans isomerisation. J. Mol. Biol. 279, 449–460 (1998).

    Article  CAS  Google Scholar 

  5. Brandts, J.F., Halvorson, H.R. & Brennan, M. Consideration of the possibility that the slow step in protein denaturation reactions is due to cis-trans isomerism of proline residues. Biochemistry 14, 4953–4963 (1975).

    Article  CAS  Google Scholar 

  6. Balbach, J. & Schmid, F.X. In Mechanisms of Protein Folding (ed. Pain, R.H.) 212–237 (Oxford Univ. Press, Oxford, UK, 2000).

    Google Scholar 

  7. Kubelka, J., Eaton, W.A. & Hofrichter, J. Experimental tests of villin subdomain folding simulations. J. Mol. Biol. 329, 625–630 (2003).

    Article  CAS  Google Scholar 

  8. Myers, J.K. & Oas, T.G. Mechanisms of fast protein folding. Annu. Rev. Biochem. 71, 783–815 (2002).

    Article  CAS  Google Scholar 

  9. Schmid, F.X. Prolyl isomerases. Adv. Protein Chem. 59, 243–282 (2001).

    Article  CAS  Google Scholar 

  10. Yaffe, M.B. et al. Sequence-specific and phosphorylation-dependent proline isomerization—a potential mitotic regulatory mechanism. Science 278, 1957–1960 (1997).

    Article  CAS  Google Scholar 

  11. Fischer, G. & Aumüller, T. Regulation of peptide bond cis/trans isomerization by enzyme catalysis and its implication in physiological processes. Rev. Physiol. Biochem. Pharmacol. 148, 105–150 (2004).

    Article  Google Scholar 

  12. Lubkowski, J., Hennecke, F., Plückthun, A. & Wlodawer, A. The structural basis of phage display elucidated by the crystal structure of the N-terminal domains of G3P. Nat. Struct. Biol. 5, 140–147 (1998).

    Article  CAS  Google Scholar 

  13. Holliger, P., Riechmann, L. & Williams, R.L. Crystal structure of the two N-terminal domains of g3p from filamentous phage fd at 1.9 angström: evidence for conformational lability. J. Mol. Biol. 288, 649–657 (1999).

    Article  CAS  Google Scholar 

  14. Chatellier, J. et al. Interdomain interactions within the gene 3 protein of filamentous phage. FEBS Lett. 463, 371–374 (1999).

    Article  CAS  Google Scholar 

  15. Karlsson, F., Borrebaeck, C.A., Nilsson, N. & Malmborg-Hager, A.C. The mechanism of bacterial infection by filamentous phages involves molecular interactions between TolA and phage protein 3 domains. J. Bacteriol. 185, 2628–2634 (2003).

    Article  CAS  Google Scholar 

  16. Deng, L.W. & Perham, R.N. Delineating the site of interaction on the pIII protein of filamentous bacteriophage fd with the F-pilus of Escherichia coli. J. Mol. Biol. 319, 603–614 (2002).

    Article  CAS  Google Scholar 

  17. Click, E.M. & Webster, R.E. Filamentous phage infection: required interactions with the TolA protein. J. Bacteriol. 179, 6464–6471 (1997).

    Article  CAS  Google Scholar 

  18. Lubkowski, J., Hennecke, F., Plückthun, A. & Wlodawer, A. Filamentous phage infection: crystal structure of g3p in complex with its coreceptor, the C-terminal domain of TolA. Structure 7, 711–722 (1999).

    Article  CAS  Google Scholar 

  19. Stengele, I., Bross, P., Garces, X., Giray, J. & Rasched, I. Dissection of functional domains in phage fd adsorption protein. Discrimination between attachment and penetration sites. J. Mol. Biol. 212, 143–149 (1990).

    Article  CAS  Google Scholar 

  20. Riechmann, L. & Holliger, P. The C-terminal domain of TolA is the coreceptor for filamentous phage infection of. E. coli. Cell 90, 351–360 (1997).

    CAS  PubMed  Google Scholar 

  21. Martin, A. & Schmid, F.X. The folding mechanism of a two-domain protein: folding kinetics and domain docking of the gene-3-protein of phage fd. J. Mol. Biol. 329, 599–610 (2003).

    Article  CAS  Google Scholar 

  22. Martin, A. & Schmid, F.X. A proline switch controls folding and domain interactions in the gene-3-protein of the filamentous phage fd. J. Mol. Biol. 331, 1131–1140 (2003).

    Article  CAS  Google Scholar 

  23. Frost, L.S. In Bacterial Conjugation (ed. Clewell, D.B.) 189–221 (Plenum Press, New York, 1993).

    Book  Google Scholar 

  24. Martin, A. & Schmid, F.X. Evolutionary stabilization of the gene-3-protein of phage fd reveals the principles that govern the thermodynamic stability of two-domain proteins. J. Mol. Biol. 328, 863–875 (2003).

    Article  CAS  Google Scholar 

  25. Lu, K.P., Hanes, S.D. & Hunter, T. A human peptidyl-prolyl isomerase essential for regulation of mitosis. Nature 380, 544–547 (1996).

    Article  CAS  Google Scholar 

  26. Andreotti, A.H. Native state proline isomerization: An intrinsic molecular switch. Biochemistry 42, 9515–9524 (2003).

    Article  CAS  Google Scholar 

  27. Weiwad, M. et al. Catalysis of proline-directed protein phosphorylation by peptidyl-prolyl cis/trans isomerases. J. Mol. Biol. 339, 635–646 (2004).

    Article  CAS  Google Scholar 

  28. Lopez-Ilasaca, M. et al. Effects of FK506-binding protein 12 and FK506 on autophosphorylation of epidermal growth factor receptor. J. Biol. Chem. 273, 9430–9434 (1998).

    Article  CAS  Google Scholar 

  29. Mallis, R.J., Brazin, K.N., Fulton, D.B. & Andreotti, A.H. Structural characterization of a proline-driven conformational switch within the Itk SH2 domain. Nat. Struct. Biol. 9, 900–905 (2002).

    Article  CAS  Google Scholar 

  30. Gitti, R.K. et al. Structure of the amino-terminal core domain of the HIV-1 capsid protein. Science 273, 231–235 (1996).

    Article  CAS  Google Scholar 

  31. Krebber, C. et al. Selectively-infective phage (SIP): a mechanistic dissection of a novel in vivo selection for protein-ligand interactions. J. Mol. Biol. 268, 607–618 (1997).

    Article  CAS  Google Scholar 

  32. Sieber, V., Plückthun, A. & Schmid, F.X. Selecting proteins with improved stability by a phage-based method. Nat. Biotechnol. 16, 955–960 (1998).

    Article  CAS  Google Scholar 

  33. Koradi, R., Billeter, M. & Wüthrich, K. MOLMOL: a program for display and analysis of macromolecular structures. J. Mol. Graph. 14, 51–55 (1996).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank C. Unverzagt for help with peptide synthesis, M. Zeeb for help with NMR measurements, G. Fischer for a sample of cyclophilin 18, P. Holliger for E. coli HB2156 and C. Lehner, W. Schumann, B. Westermann and the members of our group for comments on the manuscript. This work was supported by the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie.

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  1. Laboratorium für Biochemie und Bayreuther Zentrum für Molekulare Biowissenschaften, Universität Bayreuth, Bayreuth, D-95440, Germany

    Barbara Eckert, Andreas Martin, Jochen Balbach & Franz X Schmid

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  1. Barbara Eckert
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  2. Andreas Martin
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  3. Jochen Balbach
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Correspondence to Franz X Schmid.

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Supplementary information

Supplementary Fig. 1

Additional competitive infections. (PDF 143 kb)

Supplementary Fig. 2

Binding of TolA to N1. (PDF 181 kb)

Supplementary Methods (PDF 158 kb)

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Eckert, B., Martin, A., Balbach, J. et al. Prolyl isomerization as a molecular timer in phage infection. Nat Struct Mol Biol 12, 619–623 (2005). https://doi.org/10.1038/nsmb946

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