Crystal Structure of the Human Natural Killer Cell Activating Receptor KIR2DS2 (CD158j)

Killer cell Ig-like receptors (KIRs) regulate the function of human natural killer and T cell subsets. A feature of the KIR locus is the clustering of homologous genes encoding for inhibitory and activating KIR. Inhibitory and activating KIR differ for ligand specificities and/or affinities. In particular, we show here with KIR tetramers that activating KIR2DS2 does not bind HLA-Cw3 molecules recognized by inhibitory KIR2DL2, despite 99% extracellular amino acid identity. We also report the 2.3-Å structure of KIR2DS2, which reveals subtle displacements of two residues (Tyr45 and Gln71) involved in the interaction of KIR2DL2 with HLA-Cw3. These results show that KIR molecules cannot tolerate any variability in their three-dimensional structure without altering their MHC class I recognition capacities. Therefore, the mode of recognition used by KIR largely differs from the conformational changes that characterize T cell receptor or NKG2D interaction with their respective ligands

Investigation of the interaction between the class I MHC-related Fc receptor and its immunoglobulin G ligand

The neonatal Fc receptor (FcRn) is structurally similar to class 1 major histocompatibility molecules. FcRn transports maternal immunoglobulin G (IgG) from ingested milk into the blood. IgG is bound at the pH of milk (pH 6.0–6.5) in the gut and released at the pH of blood (pH 7.5). We find that alteration of a histidine pair within the α3 domain of FcRn and of a nearby loop (the FcRn counterpart of the class I CD8-binding loop) affects the affinity for IgG. Inhibition studies suggest the involvement of the FcRn B_2-microglobulin domain in IgG binding. Fragment B of protein A inhibits FcRn binding to IgG, localizing the binding site on Fc for FcRn to the C_H2-C_H3 domain interface. Three histidines present at the C_H2-C_H3 domain interface of Fc could be partially responsible for the pH-dependent interaction between FcRn and IgG

Crystallization and Stoichiometry of Binding of a Complex between a Rat Intestinal Fc Receptor and Fc

Fc receptors expressed in the gut of newborn rodents bind to maternal immunoglobulin in milk at pH 6·5, and transport it to the bloodstream of the neonate, where it dissociates at pH 7·4. The rat intestinal Fc receptor (FcRn) consists of a heavy chain, with significant sequence similarity to the heavy chain of class I MHC molecules, complexed to the class I light chain, β2-microglobulin. Although FcRn is predicted to contain a groove analogous to that which serves as the MHC peptide-binding site, the immunoglobulin ligand of FcRn is a macromolecule instead of a peptide. We have expressed and crystallized a secreted form of FcRn, and here report the crystallization of a complex between FcRn and its Fc ligand. Isolated FcRn-Fc complexes crystallize in space group I222 or I 212121 with unit cell dimensions a =125 Å, b=152 Å and c=216 Å. The crystals diffract to 5·5 Å resolution with anisotropic diffraction to 3·5 Å. Data collection from cryopreserved cystals may allow the resolution limit to be extended, since the major reason for the poor resolution appears to be radiation decay. Even a low-resolution view of how FcRn binds Fc would be of interest to see if the binding site corresponds to the functional part of an MHC molecule. Since the structure of Fc is known, and a structure determination of FcRn is underway, it may be possible to locate the Fc binding site on FcRn at low resolution. As an initial characterization of the FcRn-Fc mode of interaction, and to facilitate the structure determination, we have determined the stoichiometry of binding of FcRn to Fc. We show that two FcRn molecules bind per Fc, as determined by analysis of gels of washed crystals, a column binding assay, and isothermal titration calorimetry

Crystal structure at 2.2 Å resolution of the MHC-related neonatal Fc receptor

The three-dimensional structure of the rat neonatal Fc receptor (FcRn) is similar to the structure of molecules of the major histocompatibility complex (MHC). The counterpart of the MHC peptide-binding site is closed in FcRn, making the FcRn groove incapable of binding peptides. A dimer of FcRn heterodimers seen in the crystals may represent a receptor dimer that forms when the Fc portion of a single immunoglobulin binds. An alternative use of the MHC fold for immune recognition is indicated by the FcRn and FcRn/Fc co-crystal structures

Bovine α1,3-galactosyltransferase catalytic domain structure and its relationship with ABO histo-blood group and glycosphingolipid glycosyltransferases

α1,3-galactosyltransferase (α3GalT, EC 2.4.1.151) is a Golgi-resident, type II transmembrane protein that transfers galactose from UDP-α-galactose to the terminal N-acetyllactosamine unit of glycoconjugate glycans, producing the Galα1,3Galβ1,4GlcNAc oligosaccharide structure present in most mammalian glycoproteins. Unlike most other mammals, humans and Old World primates do not possess α3GalT activity, which is relevant for the hyperacute rejection observed in pig-to-human xenotransplantation. The crystal structure of the catalytic domain of substrate-free bovine α3GalT, solved and refined to 2.3 Å resolution, has a globular shape with an α/β fold containing a narrow cleft on one face, and shares a UDP-binding domain (UBD) with the recently solved inverting glycosyltransferases. The substrate-bound complex, solved and refined to 2.5 Å, allows the description of residues interacting directly with UDP-galactose. These structural data suggest that the strictly conserved residue E317 is likely to be the catalytic nucleophile involved in galactose transfer with retention of anomeric configuration as accomplished by this enzyme. Moreover, the α3GalT structure helps to identify amino acid residues that determine the specificities of the highly homologous ABO histo-blood group and glycosphingolipid glycosyltransferases

Crystal Structure of Hemolin: A Horseshoe Shape with Implications for Homophilic Adhesion

Hemolin, an insect immunoglobulin superfamily member, is a lipopolysaccharide-binding immune protein induced during bacterial infection. The 3.1 angstrom crystal structure reveals a bound phosphate and patches of positive charge, which may represent the lipopolysaccharide binding site, and a new and unexpected arrangement of four immunoglobulin-like domains forming a horseshoe. Sequence analysis and analytical ultracentrifugation suggest that the domain arrangement is a feature of the L1 family of neural cell adhesion molecules related to hemolin. These results are relevant to interpretation of human L1 mutations in neurological diseases and suggest a domain swapping model for how L1 family proteins mediate homophilic adhesion