Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Imperfect coordination chemistry facilitates metal ion release in the Psa permease

Abstract

The relative stability of divalent first-row transition metal ion complexes, as defined by the Irving-Williams series, poses a fundamental chemical challenge for selectivity in bacterial metal ion acquisition. Here we show that although the substrate-binding protein of Streptococcus pneumoniae, PsaA, is finely attuned to bind its physiological substrate manganese, it can also bind a broad range of other divalent transition metal cations. By combining high-resolution structural data, metal-binding assays and mutational analyses, we show that the inability of open-state PsaA to satisfy the preferred coordination chemistry of manganese enables the protein to undergo the conformational changes required for cargo release to the Psa permease. This is specific for manganese ions, whereas zinc ions remain bound to PsaA. Collectively, these findings suggest a new ligand binding and release mechanism for PsaA and related substrate-binding proteins that facilitate specificity for divalent cations during competition from zinc ions, which are more abundant in biological systems.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Metal ion interaction with PsaA and accumulation by S. pneumoniae.
Figure 2: The structure of open, metal-free PsaA reveals an open metal-binding site.
Figure 3: Closing of PsaA is initiated by the interaction between Asp280 and manganese.
Figure 4: Location of introduced cysteine residues in Cys-PsaA and metal ion binding by the Cys-PsaA protein.
Figure 5: Proposed metal-binding mechanism for PsaA.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank

References

  1. Papp-Wallace, K.M. & Maguire, M.E. Manganese transport and the role of manganese in virulence. Annu. Rev. Microbiol. 60, 187–209 (2006).

    Article  CAS  PubMed  Google Scholar 

  2. Gat, O. et al. The solute-binding component of a putative Mn(ii) ABC transporter (MntA) is a novel Bacillus anthracis virulence determinant. Mol. Microbiol. 58, 533–551 (2005).

    Article  CAS  PubMed  Google Scholar 

  3. Berry, A.M. & Paton, J.C. Sequence heterogeneity of PsaA, a 37-kilodalton putative adhesin essential for virulence of Streptococcus pneumoniae. Infect. Immun. 64, 5255–5262 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Yesilkaya, H. et al. Role of manganese-containing superoxide dismutase in oxidative stress and virulence of Streptococcus pneumoniae. Infect. Immun. 68, 2819–2826 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Janulczyk, R., Ricci, S. & Bjorck, L. MtsABC is important for manganese and iron transport, oxidative stress resistance, and virulence of Streptococcus pyogenes. Infect. Immun. 71, 2656–2664 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Horsburgh, M.J. et al. MntR modulates expression of the PerR regulon and superoxide resistance in Staphylococcus aureus through control of manganese uptake. Mol. Microbiol. 44, 1269–1286 (2002).

    Article  CAS  PubMed  Google Scholar 

  7. Ogunniyi, A.D. et al. Central role of manganese in regulation of stress responses, physiology, and metabolism in Streptococcus pneumoniae. J. Bacteriol. 192, 4489–4497 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Archibald, F.S. & Fridovich, I. Manganese and defenses against oxygen toxicity in Lactobacillus plantarum. J. Bacteriol. 145, 442–451 (1981).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Sobota, J.M. & Imlay, J.A. Iron enzyme ribulose-5-phosphate 3-epimerase in Escherichia coli is rapidly damaged by hydrogen peroxide but can be protected by manganese. Proc. Natl. Acad. Sci. USA 108, 5402–5407 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Aguirre, J.D. & Culotta, V.C. Battles with iron: manganese in oxidative stress protection. J. Biol. Chem. 287, 13541–13548 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Wu, H.J. et al. Manganese regulation of virulence factors and oxidative stress resistance in Neisseria gonorrhoeae. J. Proteomics 73, 899–916 (2010).

    Article  CAS  PubMed  Google Scholar 

  12. Keele, B.B. Jr., McCord, J.M. & Fridovich, I. Superoxide dismutase from Escherichia coli B. A new manganese-containing enzyme. J. Biol. Chem. 245, 6176–6181 (1970).

    CAS  PubMed  Google Scholar 

  13. Andreini, C., Bertini, I., Cavallaro, G., Holliday, G.L. & Thornton, J.M. Metal ions in biological catalysis: from enzyme databases to general principles. J. Biol. Inorg. Chem. 13, 1205–1218 (2008).

    Article  CAS  PubMed  Google Scholar 

  14. Dintilhac, A., Alloing, G., Granadel, C. & Claverys, J.P. Competence and virulence of Streptococcus pneumoniae: Adc and PsaA mutants exhibit a requirement for Zn and Mn resulting from inactivation of putative ABC metal permeases. Mol. Microbiol. 25, 727–739 (1997).

    Article  CAS  PubMed  Google Scholar 

  15. McAllister, L.J. et al. Molecular analysis of the psa permease complex of Streptococcus pneumoniae. Mol. Microbiol. 53, 889–901 (2004).

    Article  CAS  PubMed  Google Scholar 

  16. Marra, A., Lawson, S., Asundi, J.S., Brigham, D. & Hromockyj, A.E. In vivo characterization of the psa genes from Streptococcus pneumoniae in multiple models of infection. Microbiology 148, 1483–1491 (2002).

    Article  CAS  PubMed  Google Scholar 

  17. McDevitt, C.A. et al. A molecular mechanism for bacterial susceptibility to zinc. PLoS Pathog. 7, e1002357 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Lawrence, M.C. et al. The crystal structure of pneumococcal surface antigen PsaA reveals a metal-binding site and a novel structure for a putative ABC-type binding protein. Structure 6, 1553–1561 (1998).

    Article  CAS  PubMed  Google Scholar 

  19. Berntsson, R.P., Smits, S.H., Schmitt, L., Slotboom, D.J. & Poolman, B. A structural classification of substrate-binding proteins. FEBS Lett. 584, 2606–2617 (2010).

    Article  CAS  PubMed  Google Scholar 

  20. Woo, J.S., Zeltina, A., Goetz, B.A. & Locher, K.P. X-ray structure of the Yersinia pestis heme transporter HmuUV. Nat. Struct. Mol. Biol. 19, 1310–1315 (2012).

    Article  CAS  PubMed  Google Scholar 

  21. Korkhov, V.M., Mireku, S.A. & Locher, K.P. Structure of AMP-PNP–bound vitamin B12 transporter BtuCD-F. Nature 490, 367–372 (2012).

    Article  CAS  PubMed  Google Scholar 

  22. Quiocho, F.A. & Ledvina, P.S. Atomic structure and specificity of bacterial periplasmic receptors for active transport and chemotaxis: variation of common themes. Mol. Microbiol. 20, 17–25 (1996).

    Article  CAS  PubMed  Google Scholar 

  23. Mao, B., Pear, M.R., McCammon, J.A. & Quiocho, F.A. Hinge-bending in L-arabinose-binding protein. The “Venus's-flytrap” model. J. Biol. Chem. 257, 1131–1133 (1982).

    CAS  PubMed  Google Scholar 

  24. Felder, C.B., Graul, R.C., Lee, A.Y., Merkle, H.P. & Sadee, W. The Venus flytrap of periplasmic binding proteins: an ancient protein module present in multiple drug receptors. AAPS PharmSci 1, E2 (1999).

    Article  CAS  PubMed  Google Scholar 

  25. Lee, Y.H., Deka, R.K., Norgard, M.V., Radolf, J.D. & Hasemann, C.A. Treponema pallidum TroA is a periplasmic zinc-binding protein with a helical backbone. Nat. Struct. Biol. 6, 628–633 (1999).

    Article  CAS  PubMed  Google Scholar 

  26. Ilari, A., Alaleona, F., Petrarca, P., Battistoni, A. & Chiancone, E. The X-ray structure of the zinc transporter ZnuA from Salmonella enterica discloses a unique triad of zinc-coordinating histidines. J. Mol. Biol. 409, 630–641 (2011).

    Article  CAS  PubMed  Google Scholar 

  27. Wei, B., Randich, A.M., Bhattacharyya-Pakrasi, M., Pakrasi, H.B. & Smith, T.J. Possible regulatory role for the histidine-rich loop in the zinc transport protein, ZnuA. Biochemistry 46, 8734–8743 (2007).

    Article  CAS  PubMed  Google Scholar 

  28. Yatsunyk, L.A. et al. Structure and metal binding properties of ZnuA, a periplasmic zinc transporter from Escherichia coli. J. Biol. Inorg. Chem. 13, 271–288 (2008).

    Article  CAS  PubMed  Google Scholar 

  29. Lee, Y.H. et al. The crystal structure of Zn(ii)-free Treponema pallidum TroA, a periplasmic metal-binding protein, reveals a closed conformation. J. Bacteriol. 184, 2300–2304 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Couñago, R.M., McDevitt, C.A., Ween, M.P. & Kobe, B. Prokaryotic substrate-binding proteins as targets for antimicrobial therapies. Curr. Drug Targets 13, 1400–1410 (2012).

    Article  PubMed  Google Scholar 

  31. Waldron, K.J., Rutherford, J.C., Ford, D. & Robinson, N.J. Metalloproteins and metal sensing. Nature 460, 823–830 (2009).

    Article  CAS  PubMed  Google Scholar 

  32. Ma, Z., Jacobsen, F.E. & Giedroc, D.P. Coordination chemistry of bacterial metal transport and sensing. Chem. Rev. 109, 4644–4681 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Irving, H. & Williams, R.J.P. Order of stability of metal complexes. Nature 162, 746–747 (1948).

    Article  CAS  Google Scholar 

  34. Harding, M.M. Small revisions to predicted distances around metal sites in proteins. Acta Crystallogr. D Biol. Crystallogr. 62, 678–682 (2006).

    Article  PubMed  Google Scholar 

  35. Fraústo da Silva, J.J.R.F. & Williams, R.J.P. The Biological Chemistry of the Elements: the Inorganic Chemistry of Life 2nd edn. (Oxford University Press, New York, 2001).

  36. Giedroc, D.P. & Arunkumar, A.I. Metal sensor proteins: nature's metalloregulated allosteric switches. Dalton Trans. 3107–3120 (2007).

  37. Sun, X. et al. Crystal structure and metal binding properties of the lipoprotein MtsA, responsible for iron transport in Streptococcus pyogenes. Biochemistry 48, 6184–6190 (2009).

    Article  CAS  PubMed  Google Scholar 

  38. Rukhman, V., Anati, R., Melamed-Frank, M. & Adir, N. The MntC crystal structure suggests that import of Mn2+ in cyanobacteria is redox controlled. J. Mol. Biol. 348, 961–969 (2005).

    Article  CAS  PubMed  Google Scholar 

  39. Banerjee, S., Wei, B., Bhattacharyya-Pakrasi, M., Pakrasi, H.B. & Smith, T.J. Structural determinants of metal specificity in the zinc transport protein ZnuA from Synechocystis 6803. J. Mol. Biol. 333, 1061–1069 (2003).

    Article  CAS  PubMed  Google Scholar 

  40. Huckle, J.W., Morby, A.P., Turner, J.S. & Robinson, N.J. Isolation of a prokaryotic metallothionein locus and analysis of transcriptional control by trace metal ions. Mol. Microbiol. 7, 177–187 (1993).

    Article  CAS  PubMed  Google Scholar 

  41. Cavet, J.S. et al. A nickel-cobalt–sensing ArsR-SmtB family repressor. Contributions of cytosol and effector binding sites to metal selectivity. J. Biol. Chem. 277, 38441–38448 (2002).

    Article  CAS  PubMed  Google Scholar 

  42. Turner, J.S., Glands, P.D., Samson, A.C. & Robinson, N.J. Zn2+-sensing by the cyanobacterial metallothionein repressor SmtB: different motifs mediate metal-induced protein-DNA dissociation. Nucleic Acids Res. 24, 3714–3721 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Pennella, M.A., Shokes, J.E., Cosper, N.J., Scott, R.A. & Giedroc, D.P. Structural elements of metal selectivity in metal sensor proteins. Proc. Natl. Acad. Sci. USA 100, 3713–3718 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Golynskiy, M.V., Gunderson, W.A., Hendrich, M.P. & Cohen, S.M. Metal binding studies and EPR spectroscopy of the manganese transport regulator MntR. Biochemistry 45, 15359–15372 (2006).

    Article  CAS  PubMed  Google Scholar 

  45. Mills, S.A. & Marletta, M.A. Metal binding characteristics and role of iron oxidation in the ferric uptake regulator from Escherichia coli. Biochemistry 44, 13553–13559 (2005).

    Article  CAS  PubMed  Google Scholar 

  46. Long, F., Vagin, A.A., Young, P. & Murshudov, G.N. BALBES: a molecular-replacement pipeline. Acta Crystallogr. D Biol. Crystallogr. 64, 125–132 (2008).

    Article  CAS  PubMed  Google Scholar 

  47. CCP4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  48. Langer, G., Cohen, S.X., Lamzin, V.S. & Perrakis, A. Automated macromolecular model building for X-ray crystallography using ARP/wARP version 7. Nat. Protoc. 3, 1171–1179 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Emsley, P., Lohkamp, B., Scott, W.G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  50. Pettersen, E.F. et al. UCSF Chimera—a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    Article  CAS  PubMed  Google Scholar 

  51. Lindahl, E., Hess, B. & Van Der Spoel, D. GROMACS 3.0: a package for molecular simulation and trajectory analysis. J. Mol. Model. 7, 306–317 (2001).

    Article  CAS  Google Scholar 

  52. Schmid, N. et al. Definition and testing of the GROMOS force-field versions 54A7 and 54B7. Eur. Biophys. J. 40, 843–856 (2011).

    Article  CAS  PubMed  Google Scholar 

  53. Tironi, I.G., Sperb, R., Smith, P.E. & Vangunsteren, W.F. A generalized reaction field method for molecular-dynamics simulations. J. Chem. Phys. 102, 5451–5459 (1995).

    Article  CAS  Google Scholar 

  54. Hess, B., Bekker, H., Berendsen, H.J.C. & Fraaije, J.G.E.M. LINCS: a linear constraint solver for molecular simulations. J. Comput. Chem. 18, 1463–1472 (1997).

    Article  CAS  Google Scholar 

  55. Miyamoto, S. & Kollman, P.A. Settle—an analytical version of the shake and rattle algorithm for rigid water models. J. Comput. Chem. 13, 952–962 (1992).

    CAS  Google Scholar 

  56. Feenstra, K.A., Hess, B. & Berendsen, H.J.C. Improving efficiency of large time-scale molecular dynamics simulations of hydrogen-rich systems. J. Comput. Chem. 20, 786–798 (1999).

    Article  CAS  PubMed  Google Scholar 

  57. Berendsen, H.J.C., Postma, J.P.M., Vangunsteren, W.F., Dinola, A. & Haak, J.R. Molecular-dynamics with coupling to an external bath. J. Chem. Phys. 81, 3684–3690 (1984).

    Article  CAS  Google Scholar 

  58. Humphrey, W., Dalke, A. & Schulten, K. VMD: visual molecular dynamics. J. Mol. Graph. 14, 33–38 (1996).

    CAS  PubMed  Google Scholar 

  59. Maiorov, V.N. & Crippin, G.M. Significance of root-mean-square deviation in comparing three-dimensional structures of globular proteins. J. Mol. Biol. 235, 625–634 (1994).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the Australian Research Council (ARC) grants DP0986578 to A.G.M. and DP120103957 to C.A.M. and the National Health and Medical Research Council (NHMRC) project grant 1022240 to C.A.M. and program grant 565526 to J.C.P., A.G.M. and B.K. M.L.O. holds an ARC Discovery Early Career Researcher Award. B.K. is a NHMRC Senior Research Fellow. J.C.P. is a NHMRC Senior Principal Research Fellow. We thank A.E. Mark for discussions.

Author information

Authors and Affiliations

Authors

Contributions

R.M.C. designed and executed the crystallographic experiments. M.P.W., S.L.B. and C.A.M. designed and executed all of the biochemical studies. M.B., J.Z. and M.L.O. executed the molecular dynamics experiments. R.M.C., M.P.W., M.L.O. and C.A.M. drafted the manuscript. All of the authors contributed to the design, analysis, discussion of the research and writing of the final manuscript.

Corresponding authors

Correspondence to Bostjan Kobe or Christopher A McDevitt.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Results, Supplementary Tables 1–4, Supplementary Movie Legend and Supplementary Figures 1–6. (PDF 4603 kb)

Supplementary Video

The movie shows the conformational changes in PsaA as the cognate physiological ligand Mn2+ binds the protein, as revealed by the crystal structures of metal-free, open-state PsaA (PDB code 3ZK7); open-state Mn2+-PsaAD280N (PDB code 3ZKA); and closed-state Mn2+-PsaA (PDB code 3ZTT). Changes in secondary structure and backbone hydrogen bonding that occur in the flexible ('spring') helix of PsaA during binding are highlighted. The movie also shows the conformational differences between closed-state Mn2+-PsaA (PDB code 3ZTT) and closed-state Zn2+-PsaA (PDB code 1PSZ) (MOV 22844 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Couñago, R., Ween, M., Begg, S. et al. Imperfect coordination chemistry facilitates metal ion release in the Psa permease. Nat Chem Biol 10, 35–41 (2014). https://doi.org/10.1038/nchembio.1382

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.1382

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing