Biochemical and functional characterization of soluble multivalent MHC L(d)/Fcγ1 and L(d)/Fcμ chimeric proteins loaded with specific peptides

Denise M. Lepley, William E. Gillanders, Nancy B. Myers, Ruth A. Robinson, Kirk W. Beisel, James L. Wisecarver, Samuel J. Pirruccello, David R. Lee, Ted H. Hansen, Ronald J. Rubocki

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

Central to the specificity of the immune system is the interaction between the T cell receptor and the major histocompatibility complex (MHC)- peptide ligand complex. To better understand the nature of this interaction, and to investigate possible avenues for specific therapeutic intervention, we have produced soluble recombinant molecules that can modulate antigen- specific T cells. Our approach involved the construction of recombinant murine genes composed of the MHC class I gene H.2L(d) and the Fc portion of immunoglobulin (Ig) heavy chain genes μ or γ1. Stable transfectants of these L(d)/Fcγ1 and L(d)/Fcμ genes generated correctly spliced transcripts and were capable of secreting chimeric protein. Immunoprecipitation analyses demonstrated the presence of chimeric L(d)/Fcγ1 and L(d)/Fcμ monomers of approximately 69 kDa and 90 kDa, respectively, as well as chimeric dimers under nonreducing conditions. The capacity of L(d)/Ig molecules to bind specific peptide ligands was demonstrated using radiolabeled peptides or with monoclonal reagents that specifically identify peptide-induced conformational changes in the L(d) ligand binding site. Soluble divalent L(d)/Fcγ1 molecules were loaded with the murine cytomegalovirus-derived peptide and other L(d)-specific peptide ligands and subsequently isolated and purified. Peptide-loaded L(d)/Fcγ1 molecules were capable of inhibiting the response of class I-restricted T cells in vitro in a peptide-specific fashion. The development of soluble multivalent chimeric proteins that possess unique properties of both the MHC class I and Ig molecules provides a valuable reagent for the study of potential mechanisms of in vitro and in vivo immune modulation.

Original languageEnglish (US)
Pages (from-to)765-774
Number of pages10
JournalTransplantation
Volume63
Issue number5
DOIs
StatePublished - Mar 15 1997

ASJC Scopus subject areas

  • Transplantation

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