Mutually exclusive binding of peptide and invariant chain to major histocompatibility complex class II antigens.
The invariant chain is a membrane protein associated with the major histocompatibility complex class II antigens both intra- and extracellularly. The extracellular portion of the human invariant chain (Ii) was expressed in Escherichia coli as a fusion protein with a polyhistidine tail and purified by metal affinity chromatography. The recombinant Ii was used as a ligand to probe binding to the cell surface of Chinese hamster ovary cells stably transfected with human class II alpha and beta genes of the DR4 isotype. We show that recombinant Ii inhibits peptide loading on class II polypeptides and also the converse; the presence of peptide in the antigen groove prevents binding of fluorescein-conjugated Ii. Moreover, blocking of Ii binding by peptide did not require a transition of the class II dimers to an SDS-stable state. A monoclonal antibody, L243, known to bind to (or close to) the peptide pocket of the class II molecule likewise blocked Ii-fluorescein binding. Further, we investigated whether or not the Ii, a variety of bacterial superantigens or the CD4 molecule, have overlapping binding sites on the class II heterodimer. Of the class II ligands tested, reduced binding was detected for the Staphylococcus superantigen type SEB on cells precincubated with soluble Ii while the binding of the other ligands was either unchanged or marginally changed. These data clarify by a direct biochemical approach the binding characteristics of Ii in comparison with other class II ligands.
Ericson
M L
ML
Laboratoire d'Immunogénétique Humaine, INSERM U.396, Institut des Cordeliers, Paris, France.
Sundström
M
M
Sansom
D M
DM
Charron
D J
DJ
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000944:Antigens, Differentiation, B-Lymphocyte; D001483:Base Sequence; D001665:Binding Sites; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D000949:Histocompatibility Antigens Class II / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D018089:Superantigens / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000944:Antigens, Differentiation, B-Lymphocyte; D015699:HIV Envelope Protein gp120; D000949:Histocompatibility Antigens Class II; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D018089:Superantigens; C045084:invariant chain