Direct binding of a soluble natural killer cell inhibitory receptor to a soluble human leukocyte antigen-Cw4 class I major histocompatibility complex molecule.
Natural killer (NK) cells expressing specific p58 NK receptors are inhibited from lysing target cells that express human leukocyte antigen (HLA)-C class I major histocompatibility complex molecules. To investigate the interaction between p58 NK receptors and HLA-Cw4, the extracellular domain of the p58 NK receptor specific for HLA-Cw4 was overexpressed in Escherichia coli and refolded from purified inclusion bodies. The refolded NK receptor is a monomer in solution. It interacts specifically with HLA-Cw4, blocking the binding of a p58-Ig fusion protein to HLA-Cw4-expressing cells, but does not block the binding of a p58-Ig fusion protein specific for HLA-Cw3 to HLA-Cw3-expressing cells. The bacterially expressed extracellular domain of HLA-Cw4 heavy chain and beta2-microglobulin were refolded in the presence of a HLA-Cw4-specific peptide. Direct binding between the soluble p58 NK receptor and the soluble HLA-Cw4-peptide complex was observed by native gel electrophoresis. Titration binding assays show that soluble monomeric receptor forms a 1:1 complex with HLA-Cw4, independent of the presence of Zn2+. The formation of complexes between soluble, recombinant molecules indicates that HLA-Cw4 is sufficient for specific ligation by the NK receptor and that neither glycoprotein requires carbohydrate for the interaction.
Fan
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QR
Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, USA.
Garboczi
D N
DN
Winter
C C
CC
Wagtmann
N
N
Long
E O
EO
Wiley
D C
DC
D000595:Amino Acid Sequence; D001483:Base Sequence; D002850:Chromatography, Gel; D017931:DNA Primers; D015236:HLA-C Antigens / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D002479:Inclusion Bodies / Q000276:immunology; D007694:Killer Cells, Natural / Q000276:immunology*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D021382:Protein Sorting Signals / Q000096:biosynthesis; D011971:Receptors, Immunologic / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D054340:Receptors, KIR; D054344:Receptors, KIR2DL3; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D001613:beta 2-Microglobulin / Q000096:biosynthesis / Q000276:immunology
D017931:DNA Primers; D015236:HLA-C Antigens; C039966:HLA-C*04 antigen; C515078:KIR2DL3 protein, human; D046911:Macromolecular Substances; D021382:Protein Sorting Signals; D011971:Receptors, Immunologic; D054340:Receptors, KIR; D054344:Receptors, KIR2DL3; D011993:Recombinant Fusion Proteins; D001613:beta 2-Microglobulin