Protein Engineering, Vol. 14, No. 3, 195-200,
March 2001
© 2001 Oxford University Press
Enzymatically mediated engineering of multivalent MHC class II–peptide chimeras
Department of Microbiology, Mount Sinai School of Medicine,1 Gustave L. Levy Place, New York, NY 10029, USA
We previously reported the genetic engineering of the first soluble, bivalent major histocompatibility complex (MHC) class II–peptide ligand for T-cell receptor (TCR). This ligand binds stably and specifically to cognate T-cells and exhibits immunomodulatory effects in vitro and in vivo. The increase in valence of MHC class II–peptide ligands was shown to parallel their avidity for cognate TCRs and potency in stimulating cognate T-cells. We describe a new enzymatic method to increase the valence of MHC–peptide ligands by cross-linking the N-glycan moieties of dimeric MHC II–peptide units through a flexible, bifunctional polyethylene glycol linker. Using this method, we generated covalently stabilized tetravalent and octavalent MHC II–peptide ligands which bound stably and specifically to cognate TCR and preserved their structural integrity in blood and lymphoid organs for 72 h. Depending on the TCR/CD4 occupancy and degree of TCR/CD4 co-clustering, the multivalent MHC II–peptide ligands polarized efficiently the antigen-specific CD4+ T-cells toward type 2 cell differentiation or induced T-cell anergy and apoptosis. The enzymatically mediated engineering of multivalent MHC–peptide ligands for cognate TCRs may provide rational grounds for the development of new therapeutic agents endowed with strong modulatory effects on antigen-specific T-cells.