Introduction of germline residues improves the stability of anti-HIV mAb 2G12-IgM.
Bottom Line: We also investigated the effects of the class switch and germline substitutions on the ligand-binding properties of 2G12 and its capacity for HIV-1 neutralization.Our results demonstrate that the introduced germline residues improve the conformational and thermal stability of 2G12-IgM without altering its overall shape and ligand-binding properties.Interestingly, the engineered protein was found to exhibit much lower neutralization potency than its wild-type counterpart, indicating that potent antigen recognition is not solely responsible for IgM-mediated HIV-1 inactivation.
Affiliation: Department of Biotechnology, Vienna Institute of BioTechnology at BOKU, University of Natural Resources and Life Sciences, Vienna, Austria.Show MeSH
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Mentions: For further structural evaluation small-angle X-ray scattering (SAXS) was employed. The raw data (Fig. 5A) already allows to clearly discriminate between IgM-617 and the two 2G12-IgM variants, for which almost identical scattering curves were obtained. This is further supported by the calculated gyration radius (Rg) values showing that IgM-012 and IgM-012_GL adopt a less compact fold than IgM-617 (Fig. 5B). Notably, the determined Rg values are in good agreement with those of previous SAXS experiments on IgMs [33,34]. A five-fold symmetry was imposed for low-resolution shape modeling of IgM-012 and IgM-617, and a total of 50 structures were used to generate density models. In agreement with the scattering curves and the pair distance distribution function, the superimposition of the shape models confirms that IgM-617 adopts a more compact conformation than IgM-012 and IgM-012_GL (Fig. 5C).
Affiliation: Department of Biotechnology, Vienna Institute of BioTechnology at BOKU, University of Natural Resources and Life Sciences, Vienna, Austria.