Structure of human IgM rheumatoid factor Fab bound to its autoantigen IgG Fc reveals a novel topology of antibody—antigen interaction

Rheumatoid factors are the characteristic autoantibodies of rheumatoid arthritis, which bind to the Fc regions of IgG molecules. Here we report the crystal structure of the Fab fragment of a patient-derived IgM rheumatoid factor (RF-AN) complexed with human lgG4 Fc, at 3.2 Å resolution. This is the first structure of an autoantibody−autoantigen complex. The epitope recognised in IgG Fc includes the C2/C3 cleft region, and overlaps the binding sites of bacterial Fc-binding proteins. The antibody residues involved in autorecognition are all located at the edge of the conventional combining site surface, leaving much of the latter available, potentially, for recognition of a different antigen. Since an important contact residue is a somatic mutation, the structure implicates antigen-driven selection, following somatic mutation of germline genes, in the production of pathogenic rheumatoid factors.

1. Tighe, H. & Carson, D.A. Rheumatoid Factor. In Textbook of Rheumatology (eds. Kelley, W.N., Harris, E.D., Ruddy, S. & Sledge, C.B.) 241−249 (W.B. Saunders, Philadelphia, 1997).
2. Mannik, M., Nardella, F.A. & Sasso, E.H. Rheumatoid factors in immune complexes of patients with rheumatoid arthritis. Springer Seminars in Immunopathol. 10, 215−230 (1988). | ISI | ChemPort |
3. Zvaifler, N.J. The immunopathology of joint inflammation in rheumatoid arthritis. Adv. Immunol. 16, 265−336 (1973). | PubMed | ChemPort |
4. Vaughan, J.H. Pathogenetic concepts and origins of rheumatoid factor in rheumatoid arthritis. Arthritis Rheum. 36, 1−6 (1993). | PubMed | ISI | ChemPort |
5. Steinitz, M., Izak, G., Cohen, S., Ehrenfeld, M. & Flechner, J. Continuous production of monoclonal rheumatoid factor by EBV-transformed lymphocytes. Nature 287, 443−445 (1980). | PubMed | ISI | ChemPort |
6. Randen, I., Thompson, K.M., Natvig, J.B., Forre, O. & Waalen, K. Human monoclonal rheumatoid factors derived from the polyclonal repertoire of rheumatoid synovial tissue: production and characterisation. Clin. Exp. Immunol. 78, 13−18 (1989). | PubMed | ISI | ChemPort |
7. Brown, C.M.S., Plater-Zyberk, C., Mageed, R.A., Jefferis, R. & Maini, R.N. Analysis of immunoglobulins secreted by hybridomas derived from rheumatoid synovia. Clin. Exp. Immunol. 80, 366−372 (1990). | PubMed | ISI | ChemPort |
8. Chen, P.P. & Carson, D.A. New insights on the physiological and pathological rheumatoid factors in humans. In Autoimmunity: Physiology and Disease (eds. Coutinho, A. & Kazatchkine, M.) 247−266 (Wiley-Liss, New York, 1994).
9. Pascual, V. et al. Nucleotide sequence analysis of rheumatoid factors and polyreactive antibodies derived from patients with rheumatoid athritis reveals diverse use of V_H and V_L gene segments and extensive variability in CDR-3. Scand. J. Immunol. 36, 349−362 (1992). | PubMed | ISI | ChemPort |
10. Victor, K.D. & Capra, J.D. Light chain variable region gene repertoire in human autoantibodies. In Autoimmunity: Physiology and Disease (eds. Coutinho, A. & Kazatchkine, M.) 19−34 (Wiley-Liss, New York, 1994).
11. Pascual, V. et al. The complete nucleotide sequences of the heavy chain variable regions of six monopecific rheumatoid factors derived from Epstein-Barr virus transformed B cells isolated from the synovial tissue of patients with rheumatoid arthritis. J. Clin. Invest. 86, 1320−1328 (1990). | PubMed | ISI | ChemPort |
12. Olee, T. et al. Genetic analysis of self-associating IgG rheumatoid factors from two rheumatoid synovia implicates an antigen-driven response. J. Exp. Med. 175, 831−842 (1992). | PubMed | ISI | ChemPort |
13. Ermel, R.W., Kenny, T.P., Chen, P.P. & Robbins, D.L. Molecular analysis of rheumatoid factors derived from rheumatoid synovium suggests an antigen-driven response in inflamed joints. Arthritis Rheum. 36, 380−388 (1993). | PubMed | ISI | ChemPort |
14. Sasso, E.H., Barber, C.V., Nardella, F.A., Yount, W.J. & Mannik, M. Antigenic specificities of human monoclonal and polyclonal IgM rheumatoid factors. The C2- C3 interface region contains the major determinants. J. Immunol. 140, 3098−3107 (1988). | PubMed | ISI | ChemPort |
15. Bonagura, V.R. et al., Mapping studies reveal unique epitopes on IgG recognised by rheumatoid arthritis-derived monoclonal rheumatoid factors. J. Immunol. 151, 3840−3852 (1993). | PubMed | ISI | ChemPort |
16. Steinitz, M. & Tamir, S. Human monoclonal autoimmune antibody produced in vitro: rheumatoid factor generated by Epstein-Barr virus-transformed cell line. Eur. J. Immunol. 12, 126−133 (1982). | PubMed | ISI | ChemPort |
17. Sohi, M.K. et al. Crystallization of a complex between the Fab fragment of a human immunoglobulin M (IgM) rheumatoid factor (RF-AN) and the Fc fragment of human lgG4 Fc. Immunology 88, 636−641 (1996). | Article | PubMed | ISI | ChemPort |
18. Jefferis, R., Nik Jaafer, M.I. & Steinitz, M. Immunogenic and antigenic epitopes of immunoglobulins. VIII. A human monoclonal rheumatoid factor having specificity for a discontinuous epitope determined by histidine/arginine interchange at residue 435 of immunoglobulin G. Immunol. Lett. 7, 191−194 (1984). | Article | PubMed | ISI | ChemPort |
19. Padlan, E.A. Anatomy of the antibody molecule. Mol. Immunol. 31, 169−217 (1994). | Article | PubMed | ISI | ChemPort |
20. Wilson, I.A. & Stanfield, R.L. Antibody-antigen interactions: new structures and new conformational changes. Curr. Opin. Struct. Biol. 4, 857−867 (1994). | PubMed | ISI | ChemPort |
21. Malby, R.L. et al. The structure of a complex between the NC10 antibody and influenza virus neuraminidase and comparison with the overlapping binding site of the NC41 antibody. Structure 2, 733−746 (1994). | PubMed | ISI | ChemPort |
22. Connolly, M.L. Analytical molecular surface calculation. J. Appl. Crystallogr. 16, 548−558 (1983). | Article | ISI | ChemPort |
23. Davies, D.R. & Cohen, G.H. Interactions of protein antigens with antibodies. Proc. Natl. Acad. Sci. USA 93, 7−12 (1996). | Article | PubMed | ChemPort |
24. Tulip, W.R., Varghese, J.N., Laver, W.G., Webster, R.G. & Colman, P.M. Refined crystal structure of the influenza virus N9 neuraminidase-NC41 Fab complex. J. Mol. Biol. 227, 122−148 (1992). | PubMed | ISI | ChemPort |
25. Deisenhofer, J. Crystallographic refinement and atomic models of a human Fc fragment and its complex with fragment B of protein A from Staphylococcus aureus at 2.9- and 2.8-Å resolution. Biochemistry 20, 2361−2370 (1981). | PubMed | ISI | ChemPort |
26. Sauer-Eriksson, A.E., Kleywegt, G.J., Uhlen, M. & Jones, T.A. Crystal structure of the C2 fragment of streptococcal protein G in complex with the Fc domain of human IgG. Structure 3, 265−278 (1995). | PubMed | ChemPort |
27. Burmeister, W.P., Huber, A.H. & Bjorkman, P.J. Crystal structure of the complex of rat neonatal Fc receptor with Fc. Nature 372, 379−383 (1994). | Article | PubMed | ISI | ChemPort |
28. Oppliger, I.R., Nardella, F.A., Stone, G.C. & Mannik, M. Human rheumatoid factors bear the internal image of the Fc binding region of Staphylococcal protein A. J. Exp, Med. 166, 702−710 (1987). | ISI | ChemPort |
29. Jefferis, R. et al. A comparative study of the N-linked oligosaccharide structures of human IgG subclass proteins. Biochem. J. 268, 529−537 (1990). | PubMed | ISI | ChemPort |
30. Malhotra, R. et al. Glycosylation changes of IgG associated with rheumatoid arthritis can activate complement via the mannose-binding protein. Nature Medicine 1, 237−243 (1995). | PubMed | ISI | ChemPort |
31. Parekh, R.B. et al. Galactosylation of IgG associated oligosaccharides: reduction in patients with adult and juvenile onset rheumatoid arthritis and relation to disease activity. Lancet i, 966−969 (1988). | Article |
32. Soltys, A.J. et al. The binding of synovial tissue-derived human monoclonal immunoglobulin M rheumatoid factor to immunoglobulin G preparations of differing galactose content. Scand. J. Immunol. 40, 135−143 (1994). | PubMed | ISI | ChemPort |
33. Soltys, A.J., Bond, A., Westwood, O.M.R. & Hay, F.C. The effects of altered glycosylation of IgG on rheumatoid factor-binding and immune complex formation. In Glycoimmunology (eds. Alavi, A. & Axford, J.S.) 155−160 (Plenum Press, New York, 1995). | ChemPort |
34. Newkirk, M. Fc glycosylation and rheumatoid factors. In Abnormalities of IgG Glycosylation and Immunological Disorders, (eds. Isenberg, D.A. & Rademacher, T.W.) 119−130 (John Wiley & Sons Ltd., 1996).
35. Randen, I. et al. Clonally related IgM rheumatoid factors undergo affinity maturation in rheumatoid synovial inflammation. J. Immunol. 148, 3296−3301 (1992). | PubMed | ISI | ChemPort |
36. Sohi, M.K. et al. Crystallisation and preliminary X-ray analysis of the Fab fragment of a human monoclonal IgM rheumatoid factor (2A2). J. Mol. Biol. 242, 706−708 (1994). | Article | PubMed | ISI | ChemPort |
37. Collaborative Computational Project, Number 4. The CCP4 Suite: Programs for Protein Crystallography. Acta Crystallogr., D50, 760−763 (1994).
38. Marquart, M., Deisenhofer, J., Huber, R. & Palm, W. Crystallographic refinement and atomic models of the intact immunoglobulin molecule Kol and its antigen-binding fragment at 3.0Å and 1.9Å resolution. J. Mol. Biol. 141, 369−391 (1980). | PubMed | ISI | ChemPort |
39. Navaza, J. AMoRE - an automated package for molecular replacement. Acta Crystallogr. A50, 157−163 (1994). | ChemPort |
40. Brunger, A.T. X-PLOR (Ver. 3.1): A system for Crystallography and NMR (Yale University, New Haven, CT, USA, 1992).
41. Jones, T.A., Bergdoll, M. & Kjeldgaard, M. Crystallographic Computing and Modeling Methods in Molecular Design (Springer, New York, 1993).
42. Laskowski, R.A., MacArthur, M.W., Moss, D.S. & Thornton, J.M. PROCHECK - A program to check the stereochemical quality of protein structures. J.Appl. Crystallogr. 26, 283−291 (1993). | Article | ChemPort |
43. Evans, S.V. SETOR: hardware lighted three dimensional solid model representations of macromolecules. J. Mol. Graphics 11, 134−138 (1993). | Article | ISI | ChemPort |
44. Sheriff, S. Some methods for examining the interactions between two molecules. Immunomethods 3, 191−196 (1993). | Article | ChemPort |
45. Kabat, E.A., Wu, T.T., Perry, H.M., Gottesman, K.S. & Foeler, C. Sequences of Proteins of Immunological Interest (US Department of Health and Human Services, NIH, Bethesda,MD, USA, 1991).

	Top

	Top