title: Ligands for murine NKG2D display heterogeneous binding behavior.
abstract: NKG2D transmits stimulatory signals to natural killer cells and other hematopoietic cells, leading to enhanced proliferation, cytokine secretion and target killing. Murine and human NKG2D each recognize five known class I-related molecules with distinct primary structures. Here, we used surface plasmon resonance to examine the binding of murine NKG2D to its cognate ligands: RAE-1B6 (a newly described C57BL/6J variant of RAE-1), RAE-1 delta (common to BALB and C57BL6/J), and H60 (expressed in BALB, but not C57BL/6J). While RAE-1B6 and H60 display relatively high affinities for NKG2D with K(D) in the 20-30 nM range and k(off )in the 0.03s(-1) to 0.06s(-1) range (t(1/2) approximately 10-20s); the RAE-1 delta variant binds with a lower affinity: K(D) of approximately 750 nM. Furthermore, RAE-1 delta displays biphasic kinetics with dominant k(off) of approximately 0.2s(-1) (t(1/2) approximately 3s), partially explaining the lower affinity. Thus, H60 and RAE-1B6 bind NKG2D with almost identical kinetics while sharing only 20% amino acid sequence identity; whereas other RAE-1 molecules demonstrate faster dissociation and lower affinities than RAE-1B6 despite sharing 90% sequence identity. C57BL/6J mice, although not expressing the H60 gene product, retain a high-affinity ligand for NKG2D in the form of RAE-1B6.
authors.affiliation: Barnes-Jewish Hospital, Division of Pulmonary and Critical Care Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Leonidas N
authors.initials: LN
authors.lastname: Carayannopoulos
authors.forename: Olga V
authors.initials: OV
authors.lastname: Naidenko
authors.forename: Jeremy
authors.initials: J
authors.lastname: Kinder
authors.forename: Emily L
authors.initials: EL
authors.lastname: Ho
authors.forename: Daved H
authors.initials: DH
authors.lastname: Fremont
authors.forename: Wayne  M
authors.initials: W
authors.lastname: Yokoyama
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D008024:Ligands; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D015778:Minor Histocompatibility Antigens / Q000378:metabolism*; D008969:Molecular Sequence Data; D055655:NK Cell Lectin-Like Receptor Subfamily K; D017924:Nucleopolyhedroviruses / Q000235:genetics; D011485:Protein Binding; D011971:Receptors, Immunologic / Q000378:metabolism*; D055607:Receptors, Natural Killer Cell; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D018411:Spodoptera / Q000166:cytology
chemical_list: C527085:KLRK1 protein, human; C527086:Klrk1 protein, mouse; D008024:Ligands; D008565:Membrane Proteins; D015778:Minor Histocompatibility Antigens; D055655:NK Cell Lectin-Like Receptor Subfamily K; C491220:Rae-1B6 protein, mouse; C491219:Raet1a protein, mouse; D011971:Receptors, Immunologic; D055607:Receptors, Natural Killer Cell; D011993:Recombinant Fusion Proteins; C114735:minor H antigen H60