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
Related in: MedlinePlus
Mentions: The rational design of IgM-012_GL was based on the primary sequence of IgM-012. Ten mutations were introduced into the framework of IgM-012 vH and 7 mutations into that of IgM-012 vL (Tables 1a and 1b). In essence, all residues were changed into their germline counterparts except those which are (i) known to be important for antigen binding (as identified by previous studies), (ii) located in Vernier zones, and (iii) responsible for the unique domain swap of 2G12 (Fig. 1, Tables 1a and 1b). While the percentage of identities between IgM-012 and the corresponding germline sequences is 68% (vH) and 84% (vL), these values increase to 78% and 91% in the case of IgM-012_GL, leading to an improvement of the germinality index  from 0.8 (IgM-012) to 0.9 (IgM-012_GL). However, the natural IgM-617 displays an even higher extent of homology with its germline counterpart (vH: 95%; vL: 98%). It should be noted that the mutations introduced into IgM-012 did not lead to substantial changes in hydrophobicity (IgM-012: 244 hydrophobic residues; IgM-012_GL: 243 hydrophobic residues; total number of residues in both cases: 789) and isoelectric point (IgM-012: 6.72; IgM-012_GL: 6.81).
Affiliation: Department of Biotechnology, Vienna Institute of BioTechnology at BOKU, University of Natural Resources and Life Sciences, Vienna, Austria.