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A regulatable switch mediates self-association in an immunoglobulin fold

Nature Structural & Molecular Biology volume 15, pages 965–971 (2008)Cite this article

Abstract

β-2 microglobulin (β2m) is a globular protein that self-associates into fibrillar amyloid deposits in patients undergoing hemodialysis therapy. Formation of these β-sheet–rich assemblies is a fundamental property of polypeptides that can be triggered by diverse conditions. For β2m, oligomerization into pre-amyloidogenic states occurs in specific response to coordination by Cu2+. Here we report the basis for this self-association at atomic resolution. Metal is not a direct participant in the molecular interface. Rather, binding results in distal alterations enabling the formation of two new surfaces. These interact to form a closed hexameric species. The origins of this include isomerization of a buried and conserved cis-proline previously implicated in the β2m aggregation pathway. The consequences of this isomerization are evident and reveal a molecular basis for the conversion of this robust monomeric protein into an amyloid-competent state.

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Figure 1: Model of Cu2+-dependent amyloid formation of β2m.
Figure 2: Wild-type and P32A β2m.
Figure 3: Metal-dependent aggregation.
Figure 4: Solvent-accessible surface and ribbon representations of the β2m hexamer.
Figure 5: Intersubunit contacts in the β2mholo hexamer.
Figure 6: Conformational effects of Cu2+ binding.

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References

  1. Burgess, A.W. et al. An open-and-shut case? Recent insights into the activation of EGF/ErbB receptors. Mol. Cell 12, 541–552 (2003).

    Article  CAS  PubMed  Google Scholar 

  2. Pollard, T.D. & Borisy, G.G. Cellular motility driven by assembly and disassembly of actin filaments. Cell 112, 453–465 (2003).

    Article  CAS  PubMed  Google Scholar 

  3. Egelman, E.H. & Orlova, A. Allostery, cooperativity, and different structural states in F-actin. J. Struct. Biol. 115, 159–162 (1995).

    Article  CAS  PubMed  Google Scholar 

  4. Frederick, K.B., Sept, D. & De La Cruz, E.M. Effects of solution crowding on actin polymerization reveal the energetic basis for nucleotide-dependent filament stability. J. Mol. Biol. 378, 540–550 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Sunshine, H.R., Hofrichter, J., Ferrone, F.A. & Eaton, W.A. Oxygen binding by sickle cell hemoglobin polymers. J. Mol. Biol. 158, 251–273 (1982).

    Article  CAS  PubMed  Google Scholar 

  6. Rochet, J.C. & Lansbury, P.T., Jr. Amyloid fibrillogenesis: themes and variations. Curr. Opin. Struct. Biol. 10, 60–68 (2000).

    Article  CAS  PubMed  Google Scholar 

  7. Chiti, F. & Dobson, C.M. Protein misfolding, functional amyloid, and human disease. Annu. Rev. Biochem. 75, 333–366 (2006).

    Article  CAS  PubMed  Google Scholar 

  8. Santos, M. et al. Defective iron homeostasis in beta 2-microglobulin knockout mice recapitulates hereditary hemochromatosis in man. J. Exp. Med. 184, 1975–1985 (1996).

    Article  CAS  PubMed  Google Scholar 

  9. Drake, D.R., III & Lukacher, A.E. β2-microglobulin knockout mice are highly susceptible to polyoma virus tumorigenesis. Virology 252, 275–284 (1998).

    Article  CAS  PubMed  Google Scholar 

  10. Kameda, A. et al. Nuclear magnetic resonance characterization of the refolding intermediate of β2-microglobulin trapped by non-native prolyl peptide bond. J. Mol. Biol. 348, 383–397 (2005).

    Article  CAS  PubMed  Google Scholar 

  11. Jahn, T.R., Parker, M.J., Homans, S.W. & Radford, S.E. Amyloid formation under physiological conditions proceeds via a native-like folding intermediate. Nat. Struct. Mol. Biol. 13, 195–201 (2006).

    Article  CAS  PubMed  Google Scholar 

  12. Chiti, F. et al. Detection of two partially structured species in the folding process of the amyloidogenic protein β2-microglobulin. J. Mol. Biol. 307, 379–391 (2001).

    Article  CAS  PubMed  Google Scholar 

  13. Eakin, C.M., Attenello, F.J., Morgan, C.J. & Miranker, A.D. Oligomeric assembly of native-like precursors precedes amyloid formation by β2 microglobulin. Biochemistry 43, 7808–7815 (2004).

    Article  CAS  PubMed  Google Scholar 

  14. Okon, M., Bray, P. & Vucelic, D. 1H NMR assignments and secondary structure of human β2-microglobulin in solution. Biochemistry 31, 8906–8915 (1992).

    Article  CAS  PubMed  Google Scholar 

  15. Morgan, C.J., Gelfand, M., Atreya, C. & Miranker, A.D. Kidney dialysis-associated amyloidosis: a molecular role for copper in fiber formation. J. Mol. Biol. 309, 339–345 (2001).

    Article  CAS  PubMed  Google Scholar 

  16. Floege, J. & Ehlerding, G. β2-microglobulin-associated amyloidosis. Nephron 72, 9–26 (1996).

    Article  CAS  PubMed  Google Scholar 

  17. Menaa, C., Esser, E. & Sprague, S.M. β2-microglobulin stimulates osteoclast formation. Kidney Int. 73, 1275–1281 (2008).

    Article  CAS  PubMed  Google Scholar 

  18. Yamamoto, S. et al. Low concentrations of sodium dodecyl sulfate induce the extension of β2-microglobulin-related amyloid fibrils at a neutral pH. Biochemistry 43, 11075–11082 (2004).

    Article  CAS  PubMed  Google Scholar 

  19. Yamamoto, S. et al. Glycosaminoglycans enhance the trifluoroethanol-induced extension of β2-microglobulin-related amyloid fibrils at a neutral pH. J. Am. Soc. Nephrol. 15, 126–133 (2004).

    Article  CAS  PubMed  Google Scholar 

  20. McParland, V.J. et al. Partially unfolded states of β2-microglobulin and amyloid formation in vitro. Biochemistry 39, 8735–8746 (2000).

    Article  CAS  PubMed  Google Scholar 

  21. Esposito, G. et al. Removal of the N-terminal hexapeptide from human β2-microglobulin facilitates protein aggregation and fibril formation. Protein Sci. 9, 831–845 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Sasahara, K., Yagi, H., Naiki, H. & Goto, Y. Heat-induced conversion of β2-microglobulin and hen egg-white lysozyme into amyloid fibrils. J. Mol. Biol. 372, 981–991 (2007).

    Article  CAS  PubMed  Google Scholar 

  23. Myers, S.L. et al. A systematic study of the effect of physiological factors on β2-microglobulin amyloid formation at neutral pH. Biochemistry 45, 2311–2321 (2006).

    Article  CAS  PubMed  Google Scholar 

  24. Linse, S. et al. Nucleation of protein fibrillation by nanoparticles. Proc. Natl. Acad. Sci. USA 104, 8691–8696 (2007).

    Article  CAS  PubMed  Google Scholar 

  25. Eakin, C.M., Berman, A.J. & Miranker, A.D. A native to amyloidogenic transition regulated by a backbone trigger. Nat. Struct. Mol. Biol. 13, 202–208 (2006).

    Article  CAS  PubMed  Google Scholar 

  26. Dobson, C.M. An accidental breach of a protein's natural defenses. Nat. Struct. Mol. Biol. 13, 295–297 (2006).

    Article  CAS  PubMed  Google Scholar 

  27. Smith, A.M., Jahn, T.R., Ashcroft, A.E. & Radford, S.E. Direct observation of oligomeric species formed in the early stages of amyloid fibril formation using electrospray ionisation mass spectrometry. J. Mol. Biol. 364, 9–19 (2006).

    Article  CAS  PubMed  Google Scholar 

  28. Calabrese, M.F. & Miranker, A.D. Formation of a stable oligomer of β2 microglobulin requires only transient encounter with Cu(II). J. Mol. Biol. 367, 1–7 (2007).

    Article  CAS  PubMed  Google Scholar 

  29. Antwi, K. et al. Cu(II) organizes β2-microglobulin oligomers but is released upon amyloid formation. Protein Sci. 17, 748–759 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Eakin, C.M., Knight, J.D., Morgan, C.J., Gelfand, M.A. & Miranker, A.D. Formation of a copper specific binding site in non-native states of β2-microglobulin. Biochemistry 41, 10646–10656 (2002).

    Article  CAS  PubMed  Google Scholar 

  31. Verdone, G. et al. The solution structure of human β2-microglobulin reveals the prodromes of its amyloid transition. Protein Sci. 11, 487–499 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Cole, J.L. Analysis of heterogeneous interactions. Methods Enzymol. 384, 212–232 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Bjorkman, P.J. et al. Structure of the human class I histocompatibility antigen, HLA-A2. Nature 329, 506–512 (1987).

    Article  CAS  PubMed  Google Scholar 

  34. Richardson, J.S. & Richardson, D.C. Natural β-sheet proteins use negative design to avoid edge-to-edge aggregation. Proc. Natl. Acad. Sci. USA 99, 2754–2759 (2002).

    Article  CAS  PubMed  Google Scholar 

  35. Trinh, C.H., Smith, D.P., Kalverda, A.P., Phillips, S.E. & Radford, S.E. Crystal structure of monomeric human β2-microglobulin reveals clues to its amyloidogenic properties. Proc. Natl. Acad. Sci. USA 99, 9771–9776 (2002).

    Article  CAS  PubMed  Google Scholar 

  36. Iwata, K., Matsuura, T., Sakurai, K., Nakagawa, A. & Goto, Y. High-resolution crystal structure of β2-microglobulin formed at pH 7.0. J. Biochem. 142, 413–419 (2007).

    Article  CAS  PubMed  Google Scholar 

  37. Rosano, C. et al. β2-microglobulin H31Y variant 3D structure highlights the protein natural propensity towards intermolecular aggregation. J. Mol. Biol. 335, 1051–1064 (2004).

    Article  CAS  PubMed  Google Scholar 

  38. Lim, J. & Vachet, R.W. Using mass spectrometry to study copper-protein binding under native and non-native conditions: β2-microglobulin. Anal. Chem. 76, 3498–3504 (2004).

    Article  CAS  PubMed  Google Scholar 

  39. Platt, G.W., Routledge, K.E., Homans, S.W. & Radford, S.E. Fibril growth kinetics reveal a region of β2-microglobulin important for nucleation and elongation of aggregation. J. Mol. Biol. 378, 251–263 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Dobson, C.M. Protein folding and misfolding. Nature 426, 884–890 (2003).

    Article  CAS  PubMed  Google Scholar 

  41. Iwata, K. et al. 3D structure of amyloid protofilaments of β2-microglobulin fragment probed by solid-state NMR. Proc. Natl. Acad. Sci. USA 103, 18119–18124 (2006).

    Article  CAS  PubMed  Google Scholar 

  42. Jones, S., Smith, D.P. & Radford, S.E. Role of the N and C-terminal strands of β2-microglobulin in amyloid formation at neutral pH. J. Mol. Biol. 330, 935–941 (2003).

    Article  CAS  PubMed  Google Scholar 

  43. McParland, V.J., Kalverda, A.P., Homans, S.W. & Radford, S.E. Structural properties of an amyloid precursor of β2-microglobulin. Nat. Struct. Biol. 9, 326–331 (2002).

    Article  CAS  PubMed  Google Scholar 

  44. Hoshino, M. et al. Mapping the core of the β2-microglobulin amyloid fibril by H/D exchange. Nat. Struct. Biol. 9, 332–336 (2002).

    Article  CAS  PubMed  Google Scholar 

  45. Jahn, T.R., Tennent, G.A. & Radford, S.E. A common β-sheet architecture underlies in vitro and in vivo β2-microglobulin amyloid fibrils. J. Biol. Chem. 283, 17279–17286 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Ramachander, R. & Bowie, J.U. SAM domains can utilize similar surfaces for the formation of polymers and closed oligomers. J. Mol. Biol. 342, 1353–1358 (2004).

    Article  CAS  PubMed  Google Scholar 

  47. Ivanova, M.I., Sawaya, M.R., Gingery, M., Attinger, A. & Eisenberg, D. An amyloid-forming segment of β2-microglobulin suggests a molecular model for the fibril. Proc. Natl. Acad. Sci. USA 101, 10584–10589 (2004).

    Article  CAS  PubMed  Google Scholar 

  48. Bennett, M.J., Schlunegger, M.P. & Eisenberg, D. 3D domain swapping: a mechanism for oligomer assembly. Protein Sci. 4, 2455–2468 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Wright, C.F., Teichmann, S.A., Clarke, J. & Dobson, C.M. The importance of sequence diversity in the aggregation and evolution of proteins. Nature 438, 878–881 (2005).

    Article  CAS  PubMed  Google Scholar 

  50. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  PubMed  Google Scholar 

  51. Storoni, L.C., McCoy, A.J. & Read, R.J. Likelihood-enhanced fast rotation functions. Acta Crystallogr. D Biol. Crystallogr. 60, 432–438 (2004).

    Article  PubMed  Google Scholar 

  52. Terwilliger, T.C. Maximum-likelihood density modification. Acta Crystallogr. D Biol. Crystallogr. 56, 965–972 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Collaborative Computational Project, Number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  54. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  PubMed  Google Scholar 

  55. Lovell, S.C. et al. Structure validation by Cα geometry: φ, ψ and Cα deviation. Proteins 50, 437–450 (2003).

    Article  CAS  PubMed  Google Scholar 

  56. Laskowski, R., MacArthur, M., Moss, D. & Thornton, J. PROCHECK: a program to check the stereochemical quality of protein structures. J. Appl. Crystallogr. 26, 283–291 (1993).

    Article  CAS  Google Scholar 

  57. Kleywegt, G.J. & Brunger, A.T. Checking your imagination: applications of the free R value. Structure 4, 897–904 (1996).

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

We thank D.M. Engelman, A. Schepartz and G.W. Brudvig for use of instruments and the staff of the Center for Structural Biology and National Synchotron Light Source (NSLS) beamline X12B for assistance and helpful discussions. We are also grateful to Y. Xiong, Y. Modis, A. Berman, A. Valentine, S.A. Strobel, J.C. Cochrane, C. Craig, D.V. Blaho and A.O. Olivares for assistance and comments. This work was supported by the US National Institutes of Health (DK54899 and 5T32GM07223).

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Authors and Affiliations

  1. Department of Molecular Biophysics and Biochemistry, Yale University, 260 Whitney Avenue, New Haven, 06520-8114, Connecticut, USA

    Matthew F Calabrese, Catherine M Eakin, Jimin M Wang & Andrew D Miranker

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  1. Matthew F Calabrese
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  2. Catherine M Eakin
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Contributions

M.F.C., C.M.E. and A.D.M. designed the experiments; M.F.C. conducted the experiments; M.F.C. and J.M.W. solved and refined the structure; M.F.C. and A.D.M. wrote the paper.

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Correspondence to Andrew D Miranker.

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Calabrese, M., Eakin, C., Wang, J. et al. A regulatable switch mediates self-association in an immunoglobulin fold. Nat Struct Mol Biol 15, 965–971 (2008). https://doi.org/10.1038/nsmb.1483

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