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Journal of Biomolecular NMR

, Volume 57, Issue 2, pp 167–178 | Cite as

NMR spectroscopy reveals unexpected structural variation at the protein–protein interface in MHC class I molecules

  • Monika Beerbaum
  • Martin Ballaschk
  • Natalja Erdmann
  • Christina Schnick
  • Anne Diehl
  • Barbara Uchanska-Ziegler
  • Andreas Ziegler
  • Peter SchmiederEmail author
Article

Abstract

β2-Microglobulin (β2m) is a small, monomorphic protein non-covalently bound to the heavy chain (HC) in polymorphic major histocompatibility complex (MHC) class I molecules. Given the high evolutionary conservation of structural features of β2m in various MHC molecules as shown by X-ray crystallography, β2m is often considered as a mere scaffolding protein. Using nuclear magnetic resonance (NMR) spectroscopy, we investigate here whether β2m residues at the interface to the HC exhibit changes depending on HC polymorphisms and the peptides bound to the complex in solution. First we show that human β2m can effectively be produced in deuterated form using high-cell-density-fermentation and we employ the NMR resonance assignments obtained for triple-labeled β2m bound to the HLA-B*27:09 HC to examine the β2m-HC interface. We then proceed to compare the resonances of β2m in two minimally distinct subtypes, HLA-B*27:09 and HLA-B*27:05, that are differentially associated with the spondyloarthropathy Ankylosing Spondylitis. Each of these subtypes is complexed with four distinct peptides for which structural information is already available. We find that only the resonances at the β2m-HC interface show a variation of their chemical shifts between the different complexes. This indicates the existence of an unexpected plasticity that enables β2m to accommodate changes that depend on HC polymorphism as well as on the bound peptide through subtle structural variations of the protein-protein interface.

Keywords

MHC class I HLA-B27 subtypes NMR assignment Protein flexibility Protein expression Labeling β2-Microglobulin 

Abbreviations

OD600nm

Optical density of the culture at 600 nm

MHC

Major histocompatibility complex

HLA

Human leucocyte antigen

β2m

Beta-2-microglobulin

HC

Heavy chain of an MHC class I molecule

TROSY

Transverse relaxation optimized spectroscopy

HSQC

Heteronuclear single quantum correlation

NMR

Nuclear magnetic resonance

HCDF

High cell density fermentation

BMRB

BioMagResBank

Notes

Acknowledgments

We thank Dr. B. Loll (Freie Universität Berlin) for providing us with unpublished information on β2m residues in an X-ray structure obtained at room temperature and Dr. M. Dorn for help with improving the HCDF. Support from the Leibniz-Institut für Molekulare Pharmakologie (FMP) is gratefully acknowledged. The work was funded by the Deutsche Forschungsgemeinschaft (SCHM880/9-1, UC8/2-1).

References

  1. Baker BM, Scott DR, Blevins SJ, Hawse WF (2012) Structural and dynamic control of T-cell receptor specificity, cross-reactivity, and binding mechanism. Immunol Rev 250:10–31CrossRefGoogle Scholar
  2. Bjorkman PJ, Saper MA, Samraoui B, Bennett WS, Strominger JL, Wiley DC (1987) Structure of the human class I histocompatibility antigen, HLA-A2. Nature 329:506–512ADSCrossRefGoogle Scholar
  3. Borbulevych OY, Piepenbrink KH, Gloor BE, Scott DR, Sommese RF, Cole DK, Sewell AK, Baker BM (2009) T cell receptor cross-reactivity directed by antigen-dependent tuning of peptide-MHC molecular flexibility. Immunity 31:885–896CrossRefGoogle Scholar
  4. Brown MA (2011) Progress in the genetics of ankylosing spondylitis. Brief Funct Genomics 10:249–257CrossRefGoogle Scholar
  5. Cavanagh J, Palmer AG III, Skelton NJ (1996) Protein NMR spectroscopy, principles and practice. Academic Press, San DiegoGoogle Scholar
  6. Cohen LS, Becker JM, Naider F (2010) Biosynthesis of peptide fragments of eukaryotic GPCRs in Escherichia coli by directing expression into inclusion bodies. J Pept Sci 16:213–218Google Scholar
  7. D’Amato M, Fiorillo MT, Carcassi C, Mathieu A, Zuccarelli A, Bitti PP, Tosi R, Sorrentino R (1995) Relevance of residue 116 of HLA-B27 in determining susceptibility to ankylosing spondylitis. Eur J Immunol 25:3199–3201CrossRefGoogle Scholar
  8. DeLano WL (2002) The PyMOL Molecular Graphics System. San Carlos, CA, DeLano ScientificGoogle Scholar
  9. Eichner T, Kalverda AP, Thompson GS, Homans SW, Radford SE (2011) Conformational conversion during amyloid formation at atomic resolution. Mol Cell 41:161–172CrossRefGoogle Scholar
  10. Esposito G, Ricagno S, Corazza A, Rennella E, Gumral D, Mimmi MC, Betto E, Pucillo CE, Fogolari F, Viglino P, Raimondi S, Giorgetti S, Bolognesi B, Merlini G, Stoppini M, Bolognesi M, Bellotti V (2008) The controlling roles of Trp60 and Trp95 in beta2-microglobulin function, folding and amyloid aggregation properties. J Mol Biol 378:887–897CrossRefGoogle Scholar
  11. Fabian H, Huser H, Narzi D, Misselwitz R, Loll B, Ziegler A, Böckmann RA, Uchanska-Ziegler B, Naumann D (2008) HLA-B27 subtypes differentially associated with disease exhibit conformational differences in solution. J Mol Biol 376:798–810CrossRefGoogle Scholar
  12. Fabian H, Huser H, Loll B, Ziegler A, Naumann D, Uchanska-Ziegler B (2010) HLA-B27 heavy chains distinguished by a micropolymorphism exhibit differential flexibility. Arthritis Rheum 62:978–987CrossRefGoogle Scholar
  13. Fabian H, Loll B, Huser H, Naumann D, Uchanska-Ziegler B, Ziegler A (2011) Influence of inflammation-related changes on conformational characteristics of HLA-B27 subtypes as detected by IR spectroscopy. FEBS J 278:1713–1727CrossRefGoogle Scholar
  14. Fiedler S, Knocke C, Vogt J, Oschkinat H, Diehl A (2007) HCDF as a protein labeling methodology. Genet Eng Biotechnol News 27. http://www.genengnews.com/gen-articles/hcdf-as-a-protein-labeling-methodology/2090/
  15. Fiorillo MT, Rückert C, Hülsmeyer M, Sorrentino R, Saenger W, Ziegler A, Uchanska-Ziegler B (2005) Allele-dependent similarity between viral and self-peptide presentation by HLA-B27 subtypes. J Biol Chem 280:2962–2971CrossRefGoogle Scholar
  16. Gao GF, Tormo J, Gerth UC, Wyer JR, McMichael AJ, Stuart DI, Bell JI, Jones EY, Jakobsen BK (1997) Crystal structure of the complex between human CD8alpha (alpha) and HLA-A2. Nature 387:630–634ADSCrossRefGoogle Scholar
  17. Garboczi DN, Hung DT, Wiley DC (1992) HLA-A2-peptide complexes: refolding and crystallization of molecules expressed in Escherichia coli and complexed with single antigenic peptides. Proc Natl Acad Sci USA 89:3429–3433ADSCrossRefGoogle Scholar
  18. Garcia KC, Degano M, Pease LR, Huang M, Peterson PA, Teyton L, Wilson IA (1998) Structural basis of plasticity in T cell receptor recognition of a self peptide–MHC antigen. Science 279:1166–1172ADSCrossRefGoogle Scholar
  19. Giovanni R (1961) The effects of deuterium oxide on bacteria. Mol Gen Genet 402:389–402CrossRefGoogle Scholar
  20. Goddard TD, Kneller DG (2004) SPARKY 3. University of California, San FranciscoGoogle Scholar
  21. Gras S, Kjer-Nielsen L, Chen Z, Rossjohn J, McCluskey J (2011) The structural bases of direct T-cell allorecognition: implications for T-cell-mediated transplant rejection. Immunol Cell Biol 89:388–395CrossRefGoogle Scholar
  22. Günther S, Schlundt A, Sticht J, Roske Y, Heinemann U, Wiesmüller KH, Jung G, Falk K, Rotzschke O, Freund C (2010) Bidirectional binding of invariant chain peptides to an MHC class II molecule. Proc Natl Acad Sci USA 107:22219–22224ADSCrossRefGoogle Scholar
  23. Hammer RE, Maika SD, Richardson JA, Tang JP, Taurog JD (1990) Spontaneous inflammatory disease in transgenic rats expressing HLA-B27 and human beta 2m: an animal model of HLA-B27-associated human disorders. Cell 63:1099–1112CrossRefGoogle Scholar
  24. Hare BJ, Wyss DF, Osburne MS, Kern PS, Reinherz EL, Wagner G (1999) Structure, specificity and CDR mobility of a class II restricted single-chain T-cell receptor. Nat Struct Biol 6:574–581CrossRefGoogle Scholar
  25. Hawse WF, Champion MM, Joyce MV, Hellman LM, Hossain M, Ryan V, Pierce BG, Weng Z, Baker BM (2012) Cutting edge: evidence for a dynamically driven T cell signaling mechanism. J Immunol 188:5819–5823CrossRefGoogle Scholar
  26. Hee CS, Beerbaum M, Loll B, Ballaschk M, Schmieder P, Uchanska-Ziegler B, Ziegler A (2013) Dynamics of free versus complexed beta2-microglobulin and the evolution of interfaces in MHC class I molecules. Immunogenetics 65:157–172CrossRefGoogle Scholar
  27. Hodkinson JP, Jahn TR, Radford SE, Ashcroft AE (2009) HDX-ESI-MS reveals enhanced conformational dynamics of the amyloidogenic protein beta(2)-microglobulin upon release from the MHC-1. J Am Soc Mass Spectrom 20:278–286CrossRefGoogle Scholar
  28. Horton R, Wilming L, Rand V, Lovering RC, Bruford EA, Khodiyar VK, Lush MJ, Povey S, Talbot CC Jr, Wright MW, Wain HM, Trowsdale J, Ziegler A, Beck S (2004) Gene map of the extended human MHC. Nat Rev Genetics 5:889–899CrossRefGoogle Scholar
  29. Hülsmeyer M, Hillig RC, Volz A, Ruhl M, Schröder W, Saenger W, Ziegler A, Uchanska-Ziegler B (2002) HLA-B27 subtypes differentially associated with disease exhibit subtle structural alterations. J Biol Chem 277:47844–47853CrossRefGoogle Scholar
  30. Hülsmeyer M, Fiorillo MT, Bettosini F, Sorrentino R, Saenger W, Ziegler A, Uchanska-Ziegler B (2004) Dual, HLA-B27 subtype-dependent conformation of a self-peptide. J Exp Med 199:271–281CrossRefGoogle Scholar
  31. Hülsmeyer M, Welfle K, Pöhlmann T, Misselwitz R, Alexiev U, Welfle H, Saenger W, Uchanska-Ziegler B, Ziegler A (2005) Thermodynamic and structural equivalence of two HLA-B27 subtypes complexed with a self-peptide. J Mol Biol 346:1367–1379CrossRefGoogle Scholar
  32. Insaidoo FK, Zajicek J, Baker BM (2009) A general and efficient approach for NMR studies of peptide dynamics in class I MHC peptide binding grooves. Biochemistry 48:9708–9710CrossRefGoogle Scholar
  33. Kjer-Nielsen L, Clements CS, Purcell AW, Brooks AG, Whisstock JC, Burrows SR, McCluskey J, Rossjohn J (2003) A structural basis for the selection of dominant alphabeta T cell receptors in antiviral immunity. Immunity 18:53–64CrossRefGoogle Scholar
  34. Kumar P, Vahedi-Faridi A, Saenger W, Ziegler A, Uchanska-Ziegler B (2009) Conformational changes within the HLA-A1:MAGE-A1 complex induced by binding of a recombinant antibody fragment with TCR-like specificity. Protein Sci 18:37–49Google Scholar
  35. Kushner DJ, Baker A, Dunstall TG (1999) Pharmacological uses and perspectives of heavy water and deuterated compounds. Can J Physiol Pharmacol 77:79–88CrossRefGoogle Scholar
  36. Leiting B, Marsilio F, O’Connell JF (1998) Predictable deuteration of recombinant proteins expressed in Escherichia coli. Anal Biochem 265:351–355CrossRefGoogle Scholar
  37. MacFerrin KD, Terranova MP, Schreiber SL, Verdine GL (1990) Overproduction and dissection of proteins by the expression-cassette polymerase chain reaction. Proc Natl Acad Sci USA 87:1937–1941ADSCrossRefGoogle Scholar
  38. Madden DR (1995) The three-dimensional structure of peptide–MHC complexes. Annu Rev Immunol 13:587–622CrossRefGoogle Scholar
  39. Madden DR, Gorga JC, Strominger JL, Wiley DC (1992) The three-dimensional structure of HLA-B27 at 2.1 A resolution suggests a general mechanism for tight peptide binding to MHC. Cell 70:1035–1048CrossRefGoogle Scholar
  40. Marley J, Lu M, Bracken C (2001) A method for efficient isotopic labeling of recombinant proteins. J Biomol NMR 20:71–75CrossRefGoogle Scholar
  41. Michielin O, Karplus M (2002) Binding free energy differences in a TCR–peptide–MHC complex induced by a peptide mutation: a simulation analysis. J Mol Biol 324:547–569CrossRefGoogle Scholar
  42. Mittermaier A, Kay LE (2006) New tools provide new insights in NMR studies of protein dynamics. Science 312:224–228ADSCrossRefGoogle Scholar
  43. Mittermaier AK, Kay LE (2009) Observing biological dynamics at atomic resolution using NMR. Trends Biochem Sci 34:601–611CrossRefGoogle Scholar
  44. Narzi D, Becker CM, Fiorillo MT, Uchanska-Ziegler B, Ziegler A, Böckmann RA (2012) Dynamical characterization of two differentially disease associated MHC class I proteins in complex with viral and self-peptides. J Mol Biol 415:429–442CrossRefGoogle Scholar
  45. Okon M, Bray P, Vucelic D (1992) 1H NMR assignments and secondary structure of human beta 2-microglobulin in solution. Biochemistry 31:8906–8915CrossRefGoogle Scholar
  46. Paliy O, Bloor D, Brockwell D, Gilbert P, Barber J (2003) Improved methods of cultivation and production of deuteriated proteins from E. coli strains grown on fully deuteriated minimal medium. J Appl Microbiol 94:580–586CrossRefGoogle Scholar
  47. Pervushin K, Riek R, Wider G, Wüthrich K (1997) Attenuated T2 relaxation by mutual cancellation of dipole-dipole coupling and chemical shift anisotropy indicates an avenue to NMR structures of very large biological macromolecules in solution. Proc Natl Acad Sci USA 94:12366–12371ADSCrossRefGoogle Scholar
  48. Pöhlmann T, Böckmann RA, Grubmüller H, Uchanska-Ziegler B, Ziegler A, Alexiev U (2004) Differential peptide dynamics is linked to major histocompatibility complex polymorphism. J Biol Chem 279:28197–28201CrossRefGoogle Scholar
  49. Reche PA, Reinherz EL (2003) Sequence variability analysis of human class I and class II MHC molecules: functional and structural correlates of amino acid polymorphisms. J Mol Biol 331:623–641CrossRefGoogle Scholar
  50. Reiser JB, Darnault C, Gregoire C, Mosser T, Mazza G, Kearney A, van der Merwe PA, Fontecilla-Camps JC, Housset D, Malissen B (2003) CDR3 loop flexibility contributes to the degeneracy of TCR recognition. Nat Immunol 4:241–247CrossRefGoogle Scholar
  51. Reveille JD (2012) Genetics of spondyloarthritis—beyond the MHC. Nat Rev Rheumatol 8:296–304CrossRefGoogle Scholar
  52. Röben M, Hahn J, Klein E, Lamparter T, Psakis G, Hughes J, Schmieder P (2010) NMR spectroscopic investigation of mobility and hydrogen bonding of the chromophore in the binding pocket of phytochrome proteins. Chemphyschem: a European journal of chemical physics and physical chemistry 11:1248–1257CrossRefGoogle Scholar
  53. Rognan D, Scapozza L, Folkers G, Daser A (1994) Molecular dynamics simulation of MHC–peptide complexes as a tool for predicting potential T cell epitopes. Biochemistry 33:11476–11485CrossRefGoogle Scholar
  54. Rückert C, Fiorillo MT, Loll B, Moretti R, Biesiadka J, Saenger W, Ziegler A, Sorrentino R, Uchanska-Ziegler B (2006) Conformational dimorphism of self-peptides and molecular mimicry in a disease-associated HLA-B27 subtype. J Biol Chem 281:2306–2316CrossRefGoogle Scholar
  55. Rudolph MG, Stanfield RL, Wilson IA (2006) How TCRs bind MHCs, peptides, and coreceptors. Annu Rev Immunol 24:419–466CrossRefGoogle Scholar
  56. Shi Y, Qi J, Iwamoto A, Gao GF (2011) Plasticity of human CD8alphaalpha binding to peptide-HLA-A*2402. Mol Immunol 48:2198–2202CrossRefGoogle Scholar
  57. Taurog JD (2009) Animal models of spondyloarthritis. Adv Exp Med Biol 649:245–254CrossRefGoogle Scholar
  58. Tugarinov V, Hwang PM, Kay LE (2004) Nuclear magnetic resonance spectroscopy of high-molecular-weight proteins. Annu Rev Biochem 73:107–146CrossRefGoogle Scholar
  59. Uchanska-Ziegler B, Ziegler A, Schmieder P (2013) Structural and dynamic features of HLA-B27 subtypes. Curr Opin Rheumatol 25:411–418Google Scholar
  60. Varani L, Bankovich AJ, Liu CW, Colf LA, Jones LL, Kranz DM, Puglisi JD, Garcia KC (2007) Solution mapping of T cell receptor docking footprints on peptide–MHC. Proc Natl Acad Sci USA 104:13080–13085ADSCrossRefGoogle Scholar
  61. Vranken WF, Boucher W, Stevens TJ, Fogh RH, Pajon A, Llinas M, Ulrich EL, Markley JL, Ionides J, Laue ED (2005) The CCPN data model for NMR spectroscopy: development of a software pipeline. Proteins 59:687–696CrossRefGoogle Scholar
  62. Wan S, Coveney P, Flower DR (2004) Large-scale molecular dynamics simulations of HLA-A*0201 complexed with a tumor-specific antigenic peptide: can the alpha3 and beta2 m domains be neglected? J Comput Chem 25:1803–1813CrossRefGoogle Scholar
  63. Winkler K, Winter A, Rueckert C, Uchanska-Ziegler B, Alexiev U (2007) Natural MHC class I polymorphism controls the pathway of peptide dissociation from HLA-B27 complexes. Biophys J 93:2743–2755CrossRefGoogle Scholar
  64. Yin Y, Mariuzza RA (2009) The multiple mechanisms of T cell receptor cross-reactivity. Immunity 31:849–851CrossRefGoogle Scholar
  65. Zacharias M, Springer S (2004) Conformational flexibility of the MHC class I alpha1–alpha2 domain in peptide bound and free states: a molecular dynamics simulation study. Biophys J 87:2203–2214CrossRefGoogle Scholar
  66. Zhu J, Rao H, Tonelli M, Westler WM, Singarapu KK, Markley JL, DeLuca HF, Assadi-Porter FM (2012) Efficient stable isotope labeling and purification of vitamin D receptor from inclusion bodies. Protein Expression Purif 85:25–31CrossRefGoogle Scholar
  67. Ziegler A, Loll B, Misselwitz R, Uchanska-Ziegler B (2009) Implications of structural and thermodynamic studies of HLA-B27 subtypes exhibiting differential association with ankylosing spondylitis. Adv Exp Med Biol 649:177–195CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Monika Beerbaum
    • 1
  • Martin Ballaschk
    • 1
  • Natalja Erdmann
    • 1
  • Christina Schnick
    • 2
  • Anne Diehl
    • 1
  • Barbara Uchanska-Ziegler
    • 2
  • Andreas Ziegler
    • 2
  • Peter Schmieder
    • 1
    Email author
  1. 1.Leibniz-Institut für Molekulare Pharmakologie (FMP)BerlinGermany
  2. 2.Institut für Immungenetik, Charité-Universitätsmedizin BerlinFreie Universität BerlinBerlinGermany

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