Masking of the Fc region in human IgG4 by constrained X-ray scattering modelling: implications for antibody function and therapy
Résumé
Of the four human IgG antibody subclasses IgG1-IgG4, IgG4 is of interest in that this does not activate complement. The solution structures of antibodies are critical to understanding function and therapeutic applications. Thus IgG4 was studied by synchrotron X-ray scattering. The Guinier X-ray radius of gyration RG increased from 5.0 nm to 5.1 nm with increase of concentration. The distance distribution function P(r) revealed a single peak at 0.3 mg/ml, which unusually became resolved into two peaks that shifted to smaller r values when the concentration increased to 1.3 mg/ml, even though the maximum dimension of IgG4 was unchanged at 17 nm. This indicated a small concentration dependence of the IgG4 solution structure. By analytical ultracentrifugation no concentration dependence in the sedimentation coefficient of 6.4 S was observed. Constrained scattering modelling resulted in solution structural determinations that showed that IgG4 has an asymmetric solution structure in which one Fab-Fc pair is closer together than the other pair, and the accessibility of one side of the Fc region is masked by the Fab regions. The averaged distances between the two Fab-Fc pairs change from 6 nm and 6 nm to more asymmetric values of 5 nm and 8 nm with increase in IgG4 concentration. We show that the averaged conformation of the Fab regions in IgG4 are able to hinder complement C1q binding to the Fc region. These results clarify IgG4 function and provide a starting point to investigate antibody stability.
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