Structural difference in the complement activation site of human IgG1 and IgG3.
The C1q binding epicentre on IgG molecules involves residues Asp(270), Lys(322), Pro(329) and Pro(331) in the C(H)2 domain. IgG1 and IgG3 are usually the most efficient of the four human IgG subclasses in activating complement and they both share all these residues. To reveal possible differences in the structural requirement for complement activation, we created a number of NIP (5-iodo-4-hydroxy-3-nitro-phenacetyl) specific IgG1 and IgG3 antibodies with parallel mutations in or near the putative C1q binding site. The mutants were tested simultaneously for antibody induced, antibody-dependent complement-mediated lysis (ADCML) at high and low antigen concentration on the target cells using sera of human, rabbit and guinea pig as complement source. In addition, we tested the antibodies against target cells decorated with the NP hapten, which has 10-fold lower affinity for the antibodies compared to the NIP hapten. We also used ELISA methods to measure complement activation. We observed a clear difference between IgG1 and IgG3 localized to residues Asp(270), Leu(334), Leu(335). For all these residues, and especially for Asp(270), IgG1 was heavily reduced in complement activation, while IgG3 was only moderated reduced, by alanine substitution. This difference was independent of the long hinge region of IgG3, demonstrated by hinge region truncation of this isotype such that it resembles that of IgG1. This report indicates the presence of structural differences between human IgG1 and IgG3 in the C1q binding site, and points to a specialization of the two isotypes with respect to complement activation.
Michaelsen
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TE
Division of Infectious Disease Control, Norwegian Institute of Public Health. terje.e.michaelsen@fhi.no
Sandlie
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I
Bratlie
D B
DB
Sandin
R H
RH
Ihle
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O
D000818:Animals; D000915:Antibody Affinity / Q000235:genetics / Q000276:immunology; D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology; D001666:Binding Sites, Antibody / Q000235:genetics / Q000276:immunology*; D003167:Complement Activation / Q000276:immunology*; D015922:Complement C1q / Q000276:immunology* / Q000378:metabolism; D006168:Guinea Pigs; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D009154:Mutation; D009592:Nitrohydroxyiodophenylacetate / Q000276:immunology; D011817:Rabbits
D007074:Immunoglobulin G; D009592:Nitrohydroxyiodophenylacetate; D015922:Complement C1q