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.

  • Perspective
  • Published:

New insights into antibody structure with implications for specificity, variable region restriction and isotype choice

Nature Reviews Immunology volume 25, pages 621–632 (2025)Cite this article

Subjects

Abstract

The mystery surrounding the mechanisms by which antibody diversity is generated was largely settled in the 1970s by the discoveries of variable gene rearrangements and somatic hypermutation. This led to the paradigm that immunoglobulins are composed of two independent domains — variable and constant — that confer specificity and effector functions, respectively. However, since these early discoveries, there have been a series of observations of communication between the variable and constant domains that affects the overall antibody structure, which suggests that immunoglobulins have a more complex, interconnected functionality than previously thought. Another unresolved issue has been the genesis of ‘restricted’ antibody responses, characterized by the use of only a few variable region gene segments, despite the enormous potential combinatorial diversity. In this Perspective, we place recent findings related to immunoglobulin structure and function in the context of these immunologically important, historically unsolved problems to propose a new model for how antibody specificity is achieved without autoreactivity.

Access through your institution
Buy or subscribe

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

Access options

Access through your institution

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

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

Fig. 1: The generation of diversity in immunoglobulins.
Fig. 2: The effect of isotype-specific sequence variations in CH1 on the overall Fab configuration.
Fig. 3: Potential functional implications of allostery between variable and constant regions of immunoglobulins related to antigen binding and class switching.

Similar content being viewed by others

References

  1. Silverstein, A. M. Splitting the difference: the germline-somatic mutation debate on generating antibody diversity. Nat. Immunol. 4, 829–833 (2003).

    Article  CAS  PubMed  Google Scholar 

  2. Sigal, N. H. & Klinman, N. R. The B-cell clonotype repertoire. Adv. Immunol. 26, 255–337 (1978).

    Article  CAS  PubMed  Google Scholar 

  3. Kreth, H. W. & Williamson, A. R. The extent of diversity of anti-hapten antibodies in inbred mice: anti-NIP (4-hydroxy-5-iodo-3-nitro-phenacetyl) antibodies in CBA/H mice. Eur. J. Immunol. 3, 141–146 (1973).

    Article  CAS  Google Scholar 

  4. Pink, J. R. L. & Askonas, B. A. Diversity of antibodies to cross-reacting nitrophenyl haptens in inbred mice. Eur. J. Immunol. 4, 426–430 (1974).

    Article  CAS  PubMed  Google Scholar 

  5. Yung, L. L. L., Cheryl Wyn‐Evans, T. & Diener, E. Ontogeny of the murine immune system: development of antigen recognition and immune responsiveness. Eur. J. Immunol. 3, 224–228 (1973).

    Article  CAS  PubMed  Google Scholar 

  6. Pasquier, L. D. Ontogeny of the immune response in animals having less than one million lymphocytes: the larvae of the toad Alytes obstetricans. Immunology 19, 353 (1970).

    PubMed  PubMed Central  Google Scholar 

  7. Leighton, P. A., Morales, J., Harriman, W. D. & Ching, K. H. V(D)J rearrangement is dispensable for producing CDR-H3 sequence diversity in a gene converting species. Front. Immunol. 9, 362694 (2018).

    Article  Google Scholar 

  8. Weber, J., Peng, H. & Rader, C. From rabbit antibody repertoires to rabbit monoclonal antibodies. Exp. Mol. Med. 49, e305–e305 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Hozumi, N. & Tonegawa, S. Evidence for somatic rearrangement of immunoglobulin genes coding for variable and constant regions. Proc. Natl Acad. Sci. USA 73, 3628–3632 (1976).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Rees, A. R. Understanding the human antibody repertoire. MAbs 12, 1729683 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  11. Saada, R., Weinberger, M., Shahaf, G. & Mehr, R. Models for antigen receptor gene rearrangement: CDR3 length. Immunol. Cell Biol. 85, 323–332 (2007).

    Article  CAS  PubMed  Google Scholar 

  12. Collins, A. M. & Jackson, K. J. L. On being the right size: antibody repertoire formation in the mouse and human. Immunogenetics 70, 143–158 (2018).

    Article  CAS  PubMed  Google Scholar 

  13. Köhler, G. & Milstein, C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256, 495–497 (1975).

    Article  PubMed  Google Scholar 

  14. Lieberman, R., Potter, M., Mushinski, E. B., Humphrey, W. & Rudikoff, S. Genetics of a new IgVH (T15 idiotype) marker in the mouse regulating natural antibody to phosphorylcholine. J. Exp. Med. 139, 983–1001 (1974).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Claflin, J. L. Uniformity in the clonal repertoire for the immune response to phosphorylcholine in mice. Eur. J. Immunol. 6, 669–674 (1976).

    Article  CAS  PubMed  Google Scholar 

  16. Winfield, J. B., Mage, R. G. & Alexander, C. B. Anti-p-azobenzenearsonate antibody of restricted heterogeneity. II. Idiotypes of antibodies produced during a 33-month period. J. Immunol. 110, 729–735 (1973).

    Article  CAS  PubMed  Google Scholar 

  17. Mage, R. G., Pincus, J. H., Alexander, C. & Freedman, M. H. Studies of hyperimmune restricted and partially restricted anti-pneumococcal polysaccharide antibodies from allotype-defined pedigreed rabbits. II. Allotypes and idiotypes in sera and electrofocused fractions. Eur. J. Immunol. 4, 560–564 (1974).

    Article  CAS  PubMed  Google Scholar 

  18. Briles, D. E. & Davie, J. M. Clonal dominance. I. Restricted nature of the IgM antibody response to group A streptococcal carbohydrate in mice. J. Exp. Med. 141, 1291–1307 (1975).

    Article  CAS  PubMed  Google Scholar 

  19. Jack, R. S., Imanishi‐Kari, T. & Rajewsky, K. Idiotypic analysis of the response of C57BL/6 mice to the (4-hydroxy-3-nitrophenyl)acetyl group. Eur. J. Immunol. 7, 559–565 (1977).

    Article  CAS  PubMed  Google Scholar 

  20. Barrett, D. J. & Ayoub, E. M. IgG2 subclass restriction of antibody to pneumococcal polysaccharides. Clin. Exp. Immunol. 63, 127 (1986).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Perlmutter, R. M., Hansburg, D., Briles, D. E., Nicolotti, R. A. & David, J. M. Subclass restriction of murine anti-carbohydrate antibodies. J. Immunol. 121, 566–572 (1978).

    Article  CAS  PubMed  Google Scholar 

  22. Briles, D. E., Forman, C., Hudak, S. & Claflin, J. L. The effects of subclass on the ability of anti-phosphocholine antibodies to protect mice from fatal infection with Streptococcus pneumoniae. J. Mol. Cell Immunol. 1, 305–309 (1984).

    CAS  PubMed  Google Scholar 

  23. Yuan, R., Casadevall, A., Spira, G. & Scharff, M. D. Isotype switching from IgG3 to IgG1 converts a nonprotective murine antibody to Cryptococcus neoformans into a protective antibody. J. Immunol. 154, 1810–1816 (1995).

    Article  CAS  PubMed  Google Scholar 

  24. Abboud, N. et al. A requirement for FcγR in antibody-mediated bacterial toxin neutralization. J. Exp. Med. 207, 2395–2405 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Saggy, I. et al. Antibody isolation from immunized animals: comparison of phage display and antibody discovery via V gene repertoire mining. Protein Eng. Des. Sel. 25, 539–549 (2012).

    Article  CAS  PubMed  Google Scholar 

  26. Latham Claflin, J., Lieberman, R. & Davie, J. M. Clonal nature of the immune response to phosphorylcholine. I. Specificity, class, and idiotype of phosphorylcholine-binding receptors on lymphoid cells. J. Exp. Med. 139, 58–73 (1974).

    Article  Google Scholar 

  27. Morahan, G., Berek, C. & Miller, J. F. A. P. An idiotypic determinant formed by both immunoglobulin constant and variable regions. Nature 301, 720–722 (1983).

    Article  CAS  PubMed  Google Scholar 

  28. Diamond, B. & Scharff, M. D. Somatic mutation of the T15 heavy chain gives rise to an antibody with autoantibody specificity. Proc. Natl Acad. Sci. USA 81, 5841–5844 (1984).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Kieber-Emmons, T. et al. The promise of the anti-idiotype concept. Front. Oncol. 2, 196 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  30. De Bono, B., Madera, M. & Chothia, C. VH gene segments in the mouse and human genomes. J. Mol. Biol. 342, 131–143 (2004).

    Article  PubMed  Google Scholar 

  31. Chevillard, C., Ozaki, J., Herring, C. D. & Riblet, R. A three-megabase yeast artificial chromosome contig spanning the C57BL mouse Igh locus. J. Immunol. 168, 5659–5666 (2002).

    Article  CAS  PubMed  Google Scholar 

  32. Larijani, M. et al. The recombination difference between mouse κ and λ segments is mediated by a pair-wise regulation mechanism. Mol. Immunol. 43, 870–881 (2006).

    Article  CAS  PubMed  Google Scholar 

  33. Thiebe, R. et al. The variable genes and gene families of the mouse immunoglobulin κ locus. Eur. J. Immunol. 29, 2072–2081 (1999).

    Article  CAS  PubMed  Google Scholar 

  34. Mikocziova, I., Greiff, V. & Sollid, L. M. Immunoglobulin germline gene variation and its impact on human disease. Genes. Immun. 22, 205–217 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Li, H., Cui, X., Pramanik, S. & Chimge, N. O. Genetic diversity of the human immunoglobulin heavy chain VH region. Immunol. Rev. 190, 53–68 (2002).

    Article  CAS  PubMed  Google Scholar 

  36. Pallarès, N., Lefebvre, S., Contet, V., Matsuda, F. & Lefranc, M. P. The human immunoglobulin heavy variable genes. Exp. Clin. Immunogenet. 16, 36–60 (1999).

    Article  PubMed  Google Scholar 

  37. Glanville, J. et al. Naive antibody gene-segment frequencies are heritable and unaltered by chronic lymphocyte ablation. Proc. Natl Acad. Sci. USA 108, 20066–20071 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Boyd, S. D. et al. Individual variation in the germline Ig gene repertoire inferred from variable region gene rearrangements. J. Immunol. 184, 6986–6992 (2010).

    Article  CAS  PubMed  Google Scholar 

  39. Choi, N. M. et al. Deep sequencing of the murine Igh repertoire reveals complex regulation of nonrandom V gene rearrangement frequencies. J. Immunol. 191, 2393–2402 (2013).

    Article  CAS  PubMed  Google Scholar 

  40. DeKosky, B. J. et al. Large-scale sequence and structural comparisons of human naive and antigen-experienced antibody repertoires. Proc. Natl Acad. Sci. USA 113, E2636–E2645 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Janeway, C. A. The immune system evolved to discriminate infectious nonself from noninfectious self. Immunol. Today 13, 11–16 (1992).

    Article  CAS  PubMed  Google Scholar 

  42. Mukherjee, J., Casadevall, A. & Scharff, M. D. Molecular characterization of the humoral responses to Cryptococcus neoformans infection and glucuronoxylomannan-tetanus toxoid conjugate immunization. J. Exp. Med. 177, 1105–1116 (1993).

    Article  CAS  PubMed  Google Scholar 

  43. Casadevall, A. & Scharff, M. D. The mouse antibody response to infection with Cryptococcus neoformans: VH and VL usage in polysaccharide binding antibodies. J. Exp. Med. 174, 151–160 (1991).

    Article  CAS  PubMed  Google Scholar 

  44. Andersen, P. S. et al. Extensive restrictions in the VH sequence usage of the human antibody response against the Rhesus D antigen. Mol. Immunol. 44, 412–422 (2007).

    Article  CAS  PubMed  Google Scholar 

  45. Adderson, E. E., Shackelford, P. G., Quinn, A. & Carroll, W. L. Restricted Ig H chain V gene usage in the human antibody response to Haemophilus influenzae type b capsular polysaccharide. J. Immunol. 147, 1667–1674 (1991).

    Article  CAS  PubMed  Google Scholar 

  46. Solin, M. L., Kaartinen, M. & Mäkelä, O. The same few V genes account for a majority of oxazolone antibodies in most mouse strains. Mol. Immunol. 29, 1357–1362 (1992).

    Article  CAS  PubMed  Google Scholar 

  47. Kocher, H. P., Berek, C. & Jaton, J. C. The immune response of BALB/c mice to phosphorylcholine is restricted to a limited number of VH- and VL-isotypes. Mol. Immunol. 18, 1027–1033 (1981).

    Article  CAS  PubMed  Google Scholar 

  48. Wysocki, L. J., Gridley, T., Huang, S., Grandea, A. G. & Gefter, M. L. Single germline VH and V κ genes encode predominating antibody variable regions elicited in strain A mice by immunization with p-azophenylarsonate. J. Exp. Med. 166, 1–11 (1987).

    Article  CAS  PubMed  Google Scholar 

  49. Williams, J. V., Weitkamp, J. H., Blum, D. L., LaFleur, B. J. & Crowe, J. E. The human neonatal B cell response to respiratory syncytial virus uses a biased antibody variable gene repertoire that lacks somatic mutations. Mol. Immunol. 47, 407 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Gorny, M. K. et al. Preferential use of the VH5-51 gene segment by the human immune response to code for antibodies against the V3 domain of HIV-1. Mol. Immunol. 46, 917 (2009).

    Article  CAS  PubMed  Google Scholar 

  51. Robbiani, D. F. et al. Convergent antibody responses to SARS-CoV-2 in convalescent individuals. Nature 584, 437–442 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Wang, Y. et al. A large-scale systematic survey reveals recurring molecular features of public antibody responses to SARS-CoV-2. Immunity 55, 1105 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Barnes, C. O. et al. SARS-CoV-2 neutralizing antibody structures inform therapeutic strategies. Nature 588, 682–687 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Barnes, C. O. et al. Structures of human antibodies bound to SARS-CoV-2 spike reveal common epitopes and recurrent features of antibodies. Cell 182, 828 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Shrestha, L. B., Tedla, N. & Bull, R. A. Broadly-neutralizing antibodies against emerging SARS-CoV-2 variants. Front. Immunol. 12, 752003 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Dejnirattisai, W. et al. The antigenic anatomy of SARS-CoV-2 receptor binding domain. Cell 184, 2183 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Rapp, M. et al. Modular basis for potent SARS-CoV-2 neutralization by a prevalent VH1-2-derived antibody class. Cell Rep. 35, 108950 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Greenspan, N. S. & Di Cera, E. Defining epitopes: it’s not as easy as it seems. Nat. Biotechnol. 17, 936–937 (1999).

    Article  CAS  PubMed  Google Scholar 

  59. Hawa, M. I. et al. Antibodies to IA-2 and GAD65 in type 1 and type 2 diabetes: isotype restriction and polyclonality. Diabetes Care 23, 228–233 (2000).

    Article  CAS  PubMed  Google Scholar 

  60. Wang, N. et al. Conserved amino acid networks involved in antibody variable domain interactions. Proteins 76, 99–114 (2009).

    Article  CAS  PubMed  Google Scholar 

  61. Scott, M. G. & Fleischman, J. B. Preferential idiotype-isotype associations in antibodies to dinitrophenyl antigens. J. Immunol. 128, 2622–2628 (1982).

    Article  CAS  PubMed  Google Scholar 

  62. Abraham, K. M. & Teale, J. M. Isotype restriction during infection of mice with the cestode Mesocestoides corti: role of immune suppression. J. Immunol. 138, 1699–1704 (1987).

    Article  CAS  PubMed  Google Scholar 

  63. Khalife, J. et al. Isotypic restriction of the antibody response to human immunodeficiency virus. AIDS Res. Hum. Retroviruses 4, 3–9 (1988).

    Article  CAS  PubMed  Google Scholar 

  64. Pritsch, O. et al. Can isotype switch modulate antigen-binding affinity and influence clonal selection? Eur. J. Immunol. 30, 3387–3395 (2000).

    Article  CAS  PubMed  Google Scholar 

  65. Eryilmaz, E. et al. Global structures of IgG isotypes expressing identical variable regions. Mol. Immunol. 56, 588–598 (2013).

    Article  CAS  PubMed  Google Scholar 

  66. Casadevall, A. & Janda, A. Immunoglobulin isotype influences affinity and specificity. Proc. Natl Acad. Sci. USA 109, 12272–12273 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Morelock, M. M. et al. Isotype choice for chimeric antibodies affects binding properties. J. Biol. Chem. 269, 13048–13055 (1994).

    Article  CAS  PubMed  Google Scholar 

  68. Tudor, D. et al. Isotype modulates epitope specificity, affinity, and antiviral activities of anti-HIV-1 human broadly neutralizing 2F5 antibody. Proc. Natl Acad. Sci. USA 109, 12680–12685 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Porter, R. R. Separation and isolation of fractions of rabbit γ-globulin containing the antibody and antigenic combining sites. Nature 182, 670–671 (1958).

    Article  CAS  PubMed  Google Scholar 

  70. Wibmer, C. K., Moore, P. L. & Morris, L. HIV broadly neutralizing antibody targets. Curr. Opin. HIV AIDS 10, 135–143 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. D’Angelo, S. et al. Many routes to an antibody heavy-chain CDR3: necessary, yet insufficient, for specific binding. Front. Immunol. 9, 395 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  72. Vandyk, L. & Meek, K. Assembly of IgH CDR3: mechanism, regulation, and influence on antibody diversity. Int. Rev. Immunol. 8, 123–133 (1992).

    Article  CAS  PubMed  Google Scholar 

  73. Xu, J. L. & Davis, M. M. Diversity in the CDR3 region of VH is sufficient for most antibody specificities. Immunity 13, 37–45 (2000).

    Article  CAS  PubMed  Google Scholar 

  74. Davis, M. M. The evolutionary and structural ‘logic’ of antigen receptor diversity. Semin. Immunol. 16, 239–243 (2004).

    Article  CAS  PubMed  Google Scholar 

  75. Chothia, C. & Lesk, A. M. Canonical structures for the hypervariable regions of immunoglobulins. J. Mol. Biol. 196, 901–917 (1987).

    Article  CAS  PubMed  Google Scholar 

  76. Gorman, J. et al. Structure of super-potent antibody CAP256-VRC26.25 in complex with HIV-1 envelope reveals a combined mode of trimer-apex recognition. Cell Rep. 31, 107488 (2020).

    Article  CAS  PubMed  Google Scholar 

  77. Liu, L. et al. An antibody class with a common CDRH3 motif broadly neutralizes sarbecoviruses. Sci. Transl. Med. 14, eabn6859 (2022).

    Article  CAS  PubMed  Google Scholar 

  78. Liu, H. et al. Human antibodies to SARS-CoV-2 with a recurring YYDRxG motif retain binding and neutralization to variants of concern including Omicron. Commun. Biol. 5, 766 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Stanfield, R. L., Zemla, A., Wilson, I. A. & Rupp, B. Antibody elbow angles are influenced by their light chain class. J. Mol. Biol. 357, 1566–1574 (2006).

    Article  CAS  PubMed  Google Scholar 

  80. Lesk, A. M. & Chothia, C. Elbow motion in the immunoglobulins involves a molecular ball-and-socket joint. Nature 335, 188–190 (1988).

    Article  CAS  PubMed  Google Scholar 

  81. Chiu, M. L., Goulet, D. R., Teplyakov, A. & Gilliland, G. L. Antibody structure and function: the basis for engineering therapeutics. Antibodies 8, 55 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  82. Toughiri, R. et al. Comparing domain interactions within antibody Fabs with κ and λ light chains. MAbs 8, 1276–1285 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Zhao, J., Zhang, B., Zhu, J., Nussinov, R. & Ma, B. Structure and energetic basis of overrepresented λ light chain in systemic light chain amyloidosis patients. Biochim. Biophys. Acta Mol. Basis Dis. 1864, 2294–2303 (2018).

    Article  CAS  PubMed  Google Scholar 

  84. Raybould, M. I. J., Turnbull, O. M., Suter, A., Guloglu, B. & Deane, C. M. Contextualising the developability risk of antibodies with λ light chains using enhanced therapeutic antibody profiling. Commun. Biol. 7, 62 (2024).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Fernández-Quintero, M. L. et al. Surprisingly fast interface and elbow angle dynamics of antigen-binding fragments. Front. Mol. Biosci. 7, 609088 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  86. Knapp, B., Dunbar, J., Alcala, M. & Deane, C. M. Variable regions of antibodies and T-cell receptors may not be sufficient in molecular simulations investigating binding. J. Chem. Theory Comput. 13, 3097–3105 (2017).

    Article  CAS  PubMed  Google Scholar 

  87. McConnell, S. A. & Casadevall, A. Immunoglobulin constant regions provide stabilization to the paratope and enforce epitope specificity. J. Biol. Chem. 300, 107397 (2024).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Röthlisberger, D., Honegger, A. & Plückthun, A. Domain interactions in the Fab fragment: a comparative evaluation of the single-chain Fv and Fab format engineered with variable domains of different stability. J. Mol. Biol. 347, 773–789 (2005).

    Article  PubMed  Google Scholar 

  89. Vidarsson, G., Dekkers, G. & Rispens, T. IgG subclasses and allotypes: from structure to effector functions. Front. Immunol. 5, 117227 (2014).

    Article  Google Scholar 

  90. Zhang, X. et al. 3D structural fluctuation of IgG1 antibody revealed by individual particle electron tomography. Sci. Rep. 5, 1–14 (2015).

    Google Scholar 

  91. Saphire, E. O., Parren, P. W. H. I., Barbas, C. F., Burton, D. R. & Wilson, I. A. Crystallization and preliminary structure determination of an intact human immunoglobulin, b12: an antibody that broadly neutralizes primary isolates of HIV-1. Acta Crystallogr. D Biol. Crystallogr. 57, 168–171 (2001).

    Article  CAS  PubMed  Google Scholar 

  92. Feige, M. J. et al. An unfolded CH1 domain controls the assembly and secretion of IgG antibodies. Mol. Cell 34, 569–579 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. Jäger, M. & Plückthun, A. Folding and assembly of an antibody Fv fragment, a heterodimer stabilized by antigen. J. Mol. Biol. 285, 2005–2019 (1999).

    Article  PubMed  Google Scholar 

  94. Feige, M. J. et al. The structural analysis of shark IgNAR antibodies reveals evolutionary principles of immunoglobulins. Proc. Natl Acad. Sci. USA 111, 8155–8160 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. Zhang, T. et al. Genetic removal of the CH1 exon enables the production of heavy chain-only IgG in mice. Front. Immunol. 9, 2202 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  96. Al Qaraghuli, M. M., Kubiak-Ossowska, K., Ferro, V. A. & Mulheran, P. A. Antibody-protein binding and conformational changes: identifying allosteric signalling pathways to engineer a better effector response. Sci. Rep. 10, 13696 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  97. Sela-Culang, I., Alon, S. & Ofran, Y. A systematic comparison of free and bound antibodies reveals binding-related conformational changes. J. Immunol. 189, 4890–4899 (2012).

    Article  CAS  PubMed  Google Scholar 

  98. Rowe, E. S. & Tanford, C. Equilibrium and kinetics of the denaturation of a homogeneous human immunoglobulin light chain. Biochemistry 12, 4822–4827 (1973).

    Article  CAS  PubMed  Google Scholar 

  99. Garber, E. & Demarest, S. J. A broad range of Fab stabilities within a host of therapeutic IgGs. Biochem. Biophys. Res. Commun. 355, 751–757 (2007).

    Article  CAS  PubMed  Google Scholar 

  100. Torres, M., Fernandez-Fuentes, N., Fiser, A. & Casadevall, A. Exchanging murine and human immunoglobulin constant chains affects the kinetics and thermodynamics of antigen binding and chimeric antibody autoreactivity. PLoS ONE 2, e1310 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  101. Ma, B., Tsai, C. J., Haliloǧlu, T. & Nussinov, R. Dynamic allostery: linkers are not merely flexible. Structure 19, 907–917 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Yogo, R. et al. The Fab portion of immunoglobulin G contributes to its binding to Fcγ receptor III. Sci. Rep. 9, 11957 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  103. Su, C. T. T., Lua, W. H., Ling, W. L. & Gan, S. K. E. Allosteric effects between the antibody constant and variable regions: a study of IgA Fc mutations on antigen binding. Antibodies 7, 20 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  104. Lua, W. H. et al. The effects of antibody engineering CH and CL in trastuzumab and pertuzumab recombinant models: impact on antibody production and antigen-binding. Sci. Rep. 8, 718 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  105. Zhao, J., Nussinov, R. & Ma, B. The allosteric effect in antibody-antigen recognition. Methods Mol. Biol. 2253, 175–183 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  106. Janda, A., Bowen, A., Greenspan, N. S. & Casadevall, A. Ig constant region effects on variable region structure and function. Front. Microbiol. 7, 22 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  107. Xia, Y., Janda, A., Eryilmaz, E., Casadevall, A. & Putterman, C. The constant region affects antigen binding of antibodies to DNA by altering secondary structure. Mol. Immunol. 56, 28–37 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  108. Bowen, A. & Casadevall, A. Revisiting the immunoglobulin intramolecular signaling hypothesis. Trends Immunol. 37, 721–723 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  109. Yang, D., Kroe-Barrett, R., Singh, S., Roberts, C. J. & Laue, T. M. IgG cooperativity — Is there allostery? Implications for antibody functions and therapeutic antibody development. MAbs 9, 1231 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. Oda, M., Kozono, H., Morii, H. & Azuma, T. Evidence of allosteric conformational changes in the antibody constant region upon antigen binding. Int. Immunol. 15, 417–426 (2003).

    Article  CAS  PubMed  Google Scholar 

  111. Pellequer, J. L., Chen, S. W., Roberts, V. A., Tainer, J. A. & Getzoff, E. D. Unraveling the effect of changes in conformation and compactness at the antibody VL-VH interface upon antigen binding. J. Mol. Recognit. 12, 267–275 (1999).

    Article  CAS  PubMed  Google Scholar 

  112. Schroeder, H. W. & Cavacini, L. Structure and function of immunoglobulins. J. Allergy Clin. Immunol. 125, S41–S52 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  113. Notkins, A. L. Polyreactivity of antibody molecules. Trends Immunol. 25, 174–179 (2004).

    Article  CAS  PubMed  Google Scholar 

  114. Kanyavuz, A., Marey-Jarossay, A., Lacroix-Desmazes, S. & Dimitrov, J. D. Breaking the law: unconventional strategies for antibody diversification. Nat. Rev. Immunol. 19, 355–368 (2019).

    Article  CAS  PubMed  Google Scholar 

  115. Prigent, J. et al. Scarcity of autoreactive human blood IgA+ memory B cells. Eur. J. Immunol. 46, 2340–2351 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  116. Planchais, C. et al. Potent human broadly SARS-CoV-2-neutralizing IgA and IgG antibodies effective against Omicron BA.1 and BA.2. J. Exp. Med. 219, e20220638 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  117. Torres, M., Fernández-Fuentes, N., Fiser, A. & Casadevall, A. The immunoglobulin heavy chain constant region affects kinetic and thermodynamic parameters of antibody variable region interactions with antigen. J. Biol. Chem. 282, 13917–13927 (2007).

    Article  CAS  PubMed  Google Scholar 

  118. Xia, Y. et al. The constant region contributes to the antigenic specificity and renal pathogenicity of murine anti-DNA antibodies. J. Autoimmun. 39, 398–411 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  119. MacCallum, R. M., Martin, A. C. R. & Thornton, J. M. Antibody-antigen interactions: contact analysis and binding site topography. J. Mol. Biol. 262, 732–745 (1996).

    Article  CAS  PubMed  Google Scholar 

  120. Vajdos, F. F. et al. Comprehensive functional maps of the antigen-binding site of an anti-ErbB2 antibody obtained with shotgun scanning mutagenesis. J. Mol. Biol. 320, 415–428 (2002).

    Article  CAS  PubMed  Google Scholar 

  121. Queen, C. et al. A humanized antibody that binds to the interleukin 2 receptor. Proc. Natl Acad. Sci. USA 86, 10029–10033 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  122. Lamminmäki, U. & Kankare, J. A. Crystal structure of a recombinant anti-estradiol Fab fragment in complex with 17β -estradiol. J. Biol. Chem. 276, 36687–36694 (2001).

    Article  PubMed  Google Scholar 

  123. Arnaout, R. et al. High-resolution description of antibody heavy-chain repertoires in humans. PLoS ONE 6, e22365 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  124. Oostindie, S. C., Lazar, G. A., Schuurman, J. & Parren, P. W. H. I. Avidity in antibody effector functions and biotherapeutic drug design. Nat. Rev. Drug Discov. 21, 715–735 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  125. Melo-Braga, M. N. et al. Unveiling the multifaceted landscape of N-glycosylation in antibody variable domains: insights and implications. Int. J. Biol. Macromol. 257, 128362 (2023).

    Article  PubMed  PubMed Central  Google Scholar 

  126. van de Bovenkamp, F. S., Hafkenscheid, L., Rispens, T. & Rombouts, Y. The emerging importance of IgG Fab glycosylation in immunity. J. Immunol. 196, 1435–1441 (2016).

    Article  PubMed  Google Scholar 

  127. Gupta, S., Jiskoot, W., Schöneich, C. & Rathore, A. S. Oxidation and deamidation of monoclonal antibody products: potential impact on stability, biological activity, and efficacy. J. Pharm. Sci. 111, 903–918 (2022).

    Article  CAS  PubMed  Google Scholar 

  128. Zhong, X. & D’Antona, A. M. A potential antibody repertoire diversification mechanism through tyrosine sulfation for biotherapeutics engineering and production. Front. Immunol. 13, 1072702 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  129. Wang, F. et al. Structural and functional characterization of glycosylation in an immunoglobulin G1 to Cryptococcus neoformans glucuronoxylomannan. Mol. Immunol. 43, 987–998 (2006).

    Article  CAS  PubMed  Google Scholar 

  130. Dimitrov, J. D. et al. A cryptic polyreactive antibody recognizes distinct clades of HIV-1 glycoprotein 120 by an identical binding mechanism. J. Biol. Chem. 289, 17767–17779 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  131. Hadzhieva, M. et al. Mechanism and functional implications of the heme-induced binding promiscuity of IgE. Biochemistry 54, 2061–2072 (2015).

    Article  CAS  PubMed  Google Scholar 

  132. Zhou, T., Hamer, D. H., Hendrickson, W. A., Sattentau, Q. J. & Kwong, P. D. Interfacial metal and antibody recognition. Proc. Natl Acad. Sci. USA 102, 14575–14580 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  133. Greenspan, N. S. & Cooper, L. J. Cooperative binding by mouse IgG3 antibodies: implications for functional affinity, effector function, and isotype restriction. Springer Semin. Immunopathol. 15, 271–291 (1993).

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, MD, USA

    Scott A. McConnell & Arturo Casadevall

Authors
  1. Scott A. McConnell
    View author publications

    Search author on:PubMed Google Scholar

  2. Arturo Casadevall
    View author publications

    Search author on:PubMed Google Scholar

Contributions

The authors contributed equally to all aspects of the article.

Corresponding author

Correspondence to Arturo Casadevall.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Reviews Immunology thanks the anonymous reviewer(s) for their contribution to the peer review of this work.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

McConnell, S.A., Casadevall, A. New insights into antibody structure with implications for specificity, variable region restriction and isotype choice. Nat Rev Immunol 25, 621–632 (2025). https://doi.org/10.1038/s41577-025-01150-9

Download citation

  • Accepted:

  • Published:

  • Issue date:

  • DOI: https://doi.org/10.1038/s41577-025-01150-9

Search

Advanced 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