title: Biophysical investigations of complement receptor 2 (CD21 and CR2)-ligand interactions reveal amino acid contacts unique to each receptor-ligand pair.
abstract: Human complement receptor type 2 (CR2 and CD21) is a cell membrane receptor, with 15 or 16 extracellular short consensus repeats (SCRs), that promotes B lymphocyte responses and bridges innate and acquired immunity. The most distally located SCRs, SCR1-2, mediate the interaction of CR2 with its four known ligands (C3d, EBV gp350, IFNalpha, and CD23). To ascertain specific interacting residues on CR2, we utilized NMR studies wherein gp350 and IFNalpha were titrated into (15)N-labeled SCR1-2, and chemical shift changes indicative of specific inter-molecular interactions were identified. With backbone assignments made, the chemical shift changes were mapped onto the crystal structure of SCR1-2. With regard to gp350, the binding region of CR2 is primarily focused on SCR1 and the inter-SCR linker, specifically residues Asn(11), Arg(13), Ala(22), Arg(28), Ser(32), Arg(36), Lys(41), Lys(57), Tyr(64), Lys(67), Tyr(68), Arg(83), Gly(84), and Arg(89). With regard to IFNalpha, the binding is similar to the CR2-C3d interaction with specific residues being Arg(13), Tyr(16), Arg(28), Ser(42), Lys(48), Lys(50), Tyr(68), Arg(83), Gly(84), and Arg(89). We also report thermodynamic properties of each ligand-receptor pair determined using isothermal titration calorimetry. The CR2-C3d interaction was characterized as a two-mode binding interaction with K(d) values of 0.13 and 160 microm, whereas the CR2-gp350 and CR2-IFNalpha interactions were characterized as single site binding events with affinities of 0.014 and 0.035 microm, respectively. The compilation of chemical binding maps suggests specific residues on CR2 that are uniquely important in each of these three binding interactions.
authors.affiliation: Department of Medicine and Immunology, University of Colorado Denver School of Medicine, Aurora, Colorado 80045.
authors.forename: James M
authors.initials: JM
authors.lastname: Kovacs
authors.affiliation: Institute of Structural and Molecular Biology, University of Edinburgh, Edinburgh EH9 3JR, Scotland, United Kingdom.
authors.forename: Jonathan P
authors.initials: JP
authors.lastname: Hannan
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Colorado Denver School of Medicine, Aurora, Colorado 80045.
authors.forename: Elan Z
authors.initials: EZ
authors.lastname: Eisenmesser
authors.affiliation: Department of Medicine and Immunology, University of Colorado Denver School of Medicine, Aurora, Colorado 80045. Electronic address: michael.holers@ucdenver.edu.
authors.forename: V Michael
authors.initials: VM
authors.lastname: Holers
mesh_terms: D056704:Adaptive Immunity / Q000502:physiology; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D015933:Complement C3d / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007113:Immunity, Innate / Q000502:physiology; D016898:Interferon-alpha / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D008024:Ligands; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding / Q000502:physiology; D020836:Protein Structure, Quaternary; D017464:Receptors, Complement 3d / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D017455:Receptors, IgE / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
chemical_list: D016898:Interferon-alpha; D008024:Ligands; D017464:Receptors, Complement 3d; D017455:Receptors, IgE; D015933:Complement C3d