ORCID Profile
0000-0003-0628-1328
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Publisher: American Chemical Society (ACS)
Date: 1996
DOI: 10.1021/BI961709E
Publisher: Elsevier BV
Date: 10-2014
DOI: 10.1016/J.FCT.2014.07.032
Abstract: Over the last 150 years a number of people in New Zealand have been incapacitated, hospitalised, or died from eating honey contaminated with tutin, a plant-derived neurotoxin. A feature of the most recent poisoning incident in 2008 was the large variability in the onset time of clinical signs and symptoms of toxicity (0.5-17 h). To investigate the basis of this variability a pharmacokinetic study was undertaken in which 6 healthy males received a single oral dose of tutin-containing honey giving a tutin dose of 1.8 μg/kg body weight. The serum concentration-time curve for all volunteers exhibited two discrete peaks with the second and higher level occurring at approximately 15 h post-dose. Two subjects reported mild, transient headache at a time post-dose corresponding to maximum tutin concentrations. There were no other signs or symptoms typical of tutin intoxication such as nausea, vomiting, dizziness or seizures. Pharmacokinetic analysis using a two-site absorption model resulted in a good fit to the observed concentration data. A novel analytical method subsequently revealed the presence of glycoside conjugates of tutin in addition to unconjugated tutin in honey. These pharmacokinetic data will be important to better define a safe maximum tutin concentration in honey.
Publisher: Elsevier BV
Date: 06-2000
Publisher: Wiley
Date: 09-1995
Abstract: The solvation of the antibody-antigen Fv D1.3-lysozyme complex is investigated through a study of the conservation of water molecules in crystal structures of the wild-type Fv fragment of antibody D1.3, 5 free lysozyme, the wild-type Fv D1.3-lysozyme complex, 5 Fv D1.3 mutants complexed with lysozyme and the crystal structure of an idiotope (Fv D1.3)-anti-idiotope (Fv E5.2) complex. In all, there are 99 water molecules common to the wild-type and mutant antibody-lysozyme complexes. The antibody-lysozyme interface includes 25 well-ordered solvent molecules, conserved among the wild-type and mutant Fv D1.3-lysozyme complexes, which are bound directly or through other water molecules to both antibody and antigen. In addition to contributing hydrogen bonds to the antibody-antigen interaction the solvent molecules fill many interface cavities. Comparison with x-ray crystal structures of free Fv D1.3 and free lysozyme shows that 20 of these conserved interface waters in the complex were bound to one of the free proteins. Up to 23 additional water molecules are also found in the antibody-antigen interface, however these waters do not bridge antibody and antigen and their temperature factors are much higher than those of the 25 well-ordered waters. Fifteen water molecules are displaced to form the complex, some of which are substituted by hydrophilic protein atoms, and 5 water molecules are added at the antibody- antigen interface with the formation of the complex. While the current crystal models of the D1.3-lysozyme complex do not demonstrate the increase in bound waters found in a physico-chemical study of the interaction at decreased water activities, the 25 well- ordered interface waters contribute a net gain of 10 hydrogen bonds to complex stability.
Publisher: Elsevier BV
Date: 12-1991
DOI: 10.1016/0022-2836(91)90593-U
Abstract: A three-dimensional model of the "blue" copper-glycoprotein stellacyanin from Rhus vernicifera has been derived by computer graphics, energy minimization and molecular dynamics techniques. The initial atomic co-ordinates were obtained by making substitutions and insertions in the known structure of another blue copper-protein, cucumber basic protein (CBP), which is 46% homologous with stellacyanin and has similar spectroscopic properties. An important difference between CBP and stellacyanin is that the latter lacks methionine, a residue that forms an exceptionally long bond to the copper atom in all blue copper-proteins of known structure. In the aligned amino acid sequences, stellacyanin has glutamine 97 at the position that corresponds to the copper-binding methionine 89 in CBP. The hypothesis that the copper atom in stellacyanin is co-ordinated by the side-chain functional groups of histidine 46, cysteine 87, histidine 92 and glutamine 97 leads to a model that enables the spectroscopic properties, redox potential and electron-transfer kinetics of the protein to be rationalized. The present model for stellacyanin is more plausible than an antecedent model derived from the structure of plastocyanin. This demonstrates that the output from molecular modeling calculations is strongly dependent on the input, and that sequence homology with the target molecule is an important criterion for the selection of a starting model.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 15-12-1995
DOI: 10.1126/SCIENCE.270.5243.1821
Abstract: The crystal structure of the V alpha domain of a T cell antigen receptor (TCR) was determined at a resolution of 2.2 angstroms. This structure represents an immunoglobulin topology set different from those previously described. A switch in a polypeptide strand from one beta sheet to the other enables a pair of V alpha homodimers to pack together to form a tetramer, such that the homodimers are parallel to each other and all hypervariable loops face in one direction. On the basis of the observed mode of V alpha association, a model of an (alpha beta)2 TCR tetramer can be positioned relative to the major histocompatibility complex class II (alpha beta)2 tetramer with the third hypervariable loop of V alpha over the amino-terminal portion of the antigenic peptide and the corresponding loop of V beta over its carboxyl-terminal residues. TCR dimerization that is mediated by the alpha chain may contribute to the coupling of antigen recognition to signal transduction during T cell activation.
Publisher: Elsevier BV
Date: 06-1994
Abstract: A recombinant form of the variable domain of the alpha chain of a murine T-cell receptor specific for the N-terminal nonapeptide of myelin basin protein in association with the major histocompatibility complex class II I-Au molecule has been crystallized in a form suitable for X-ray diffraction analysis. This protein was secreted into the periplasmic space of Escherichia coli cells and affinity-purified using a nickel chelate adsorbent. The crystals are orthorhombic, space group P2(1)2(1)2, with unit cell dimensions a = 97.7 A, b = 79.6 A, c = 30.4 A and diffract to beyond 2.2 A resolution. The ability to crystallize a T-cell receptor domain produced in bacteria strongly suggests that the periplasmic space can provide a suitable environment for the correct in vivo folding of this class of antigen recognition molecules.
Publisher: International Union of Crystallography (IUCr)
Date: 12-1992
Publisher: Proceedings of the National Academy of Sciences
Date: 05-08-1997
Abstract: Two mouse monoclonal anti-anti-idiotopic antibodies (anti-anti-Id, Ab3), AF14 and AF52, were prepared by immunizing BALB/c mice with rabbit polyclonal anti-idiotypic antibodies (anti-Id, Ab2) raised against antibody D1.3 (Ab1) specific for the antigen hen egg lysozyme. AF14 and AF52 react with an “internal image” monoclonal mouse anti-Id antibody E5.2 (Ab2), previously raised against D1.3, with affinity constants (1.0 × 10 9 M −1 and 2.4 × 10 7 M −1 , respectively) usually observed in secondary responses against protein antigens. They also react with the antigen but with lower affinity (1.8 × 10 6 M −1 and 3.8 × 10 6 M −1 ). This pattern of affinities for the anti-Id and for the antigen also was displayed by the sera of the immunized mice. The amino acid sequences of AF14 and AF52 are very close to that of D1.3. In particular, the amino acid side chains that contribute to contacts with both antigen and anti-Id are largely conserved in AF14 and AF52 compared with D1.3. Therapeutic immunizations against different pathogenic antigens using anti-Id antibodies have been proposed. Our experiments show that a response to an anti-Id immunogen elicits anti-anti-Id antibodies that are optimized for binding the anti-Id antibodies rather than the antigen.
Publisher: American Chemical Society
Date: 28-12-1990
Publisher: International Union of Crystallography (IUCr)
Date: 27-06-2003
DOI: 10.1107/S0907444903010710
Abstract: Yersinia pseudotuberculosis-derived mitogen (YPM), a superantigen with no amino-acid sequence similarity to other known superantigens, has been crystallized by the sitting-drop vapour-diffusion method. The crystals belong to space group C2, with unit-cell parameters a = 138.67, b = 78.66, c = 32.91 A, beta = 91.97 degrees. A native data set has been collected to a resolution of 1.8 A using synchrotron radiation. Self-rotation function calculations suggest the presence of three molecules in the asymmetric unit, corresponding to a solvent content of 45%.
Publisher: Springer Science and Business Media LLC
Date: 04-1995
DOI: 10.1038/374739A0
Abstract: Idiotopes are antigenic determinants, unique to an antibody or group of antibodies, defined by the reaction of anti-idiotopic antibodies with the antibodies bearing the idiotopes. The ensemble of idiotopes of an antibody constitutes its idiotype. Idiotypes are useful as markers to follow specific antibodies and clones of cells in immune responses and the inheritance of immunoglobulin genes. As external antigens and anti-idiotypic antibodies can competitively bind the combining site of specific antibodies, some anti-idiotypic antibodies may resemble the external antigen, thus mimicking its structure. It has been proposed that an anti-idiotypic antibody, anti-anti-X, may resemble the external antigen X and thus carry its 'internal image', but this idea is not unequivocally supported by the three-dimensional structures of anti-idiotopic antibodies, either because the structures of the external antigen or of the anti-idiotopic antibody were unknown, or because the anti-idiotopic antibodies showed no resemblance to the external antigens (reviewed in ref. 10). Functional mimicry of ligands of biological receptors by anti-idiotypic antibodies has been described in several systems (reviewed in ref. 11). But how closely can antibodies mimic antigens at the molecular level? Here we present the crystal structure of an idiotope-anti-idiotope complex between the Fv fragments of the anti-lysozyme antibody D1.3 and the anti-D1.3 antibody E5.2. D1.3 contacts the antigen, lysozyme and the anti-idiotopic E5.2 through essentially the same combining-site residues. In addition, E5.2 interacts with D1.3, making contacts similar to those between lysozyme and D1.3. Thus, the anti-idiotopic antibody E5.2 mimics lysozyme in its binding interactions with D1.3. Validating these observations, E5.2, used as an immunogen, induces an anti-lysozyme response.
Publisher: American Chemical Society (ACS)
Date: 22-11-2021
DOI: 10.1021/BI000704L
Abstract: Antigen-antibody complexes provide useful models for analyzing the thermodynamics of protein-protein association reactions. We have employed site-directed mutagenesis, X-ray crystallography, and isothermal titration calorimetry to investigate the role of hydrophobic interactions in stabilizing the complex between the Fv fragment of the anti-hen egg white lysozyme (HEL) antibody D1.3 and HEL. Crystal structures of six FvD1.3-HEL mutant complexes in which an interface tryptophan residue (V(L)W92) has been replaced by residues with smaller side chains (alanine, serine, valine, aspartate, histidine, and phenylalanine) were determined to resolutions between 1.75 and 2.00 A. In the wild-type complex, V(L)W92 occupies a large hydrophobic pocket on the surface of HEL and constitutes an energetic "hot spot" for antigen binding. The losses in apolar buried surface area in the mutant complexes, relative to wild-type, range from 25 (V(L)F92) to 115 A(2) (V(L)A92), with no significant shifts in the positions of protein atoms at the mutation site for any of the complexes except V(L)A92, where there is a peptide flip. The affinities of the mutant Fv fragments for HEL are 10-100-fold lower than that of the original antibody. Formation of all six mutant complexes is marked by a decrease in binding enthalpy that exceeds the decrease in binding free energy, such that the loss in enthalpy is partly offset by a compensating gain in entropy. No correlation was observed between decreases in apolar, polar, or aggregate (sum of the apolar and polar) buried surface area in the V(L)92 mutant series and changes in the enthalpy of formation. Conversely, there exist linear correlations between losses of apolar buried surface and decreases in binding free energy (R(2) = 0.937) as well as increases in the solvent portion of the entropy of binding (R(2) = 0.909). The correlation between binding free energy and apolar buried surface area corresponds to 21 cal mol(-1) A(-2) (1 cal = 4.185 J) for the effective hydrophobicity at the V(L)92 mutation site. Furthermore, the slope of the line defined by the correlation between changes in binding free energy and solvent entropy approaches unity, demonstrating that the exclusion of solvent from the binding interface is the predominant energetic factor in the formation of this protein complex. Our estimate of the hydrophobic contribution to binding at site V(L)92 in the D1.3-HEL interface is consistent with values for the hydrophobic effect derived from classical hydrocarbon solubility models. We also show how residue V(L)W92 can contribute significantly less to stabilization when buried in a more polar pocket, illustrating the dependence of the hydrophobic effect on local environment at different sites in a protein-protein interface.
Publisher: American Chemical Society
Date: 13-12-1989
Publisher: Elsevier BV
Date: 05-1994
Abstract: The three-dimensional structure of a site-directed mutant of the bacterially expressed Fv fragment from monoclonal antibody D1.3, complexed to the specific antigen lysozyme has been determined to a nominal resolution of 1.8 A using X-ray diffraction data. The replacement of VL Trp92 by Asp allows two water molecules to occupy space taken by Trp92 in the wild-type complex, in agreement with a previous observation that water molecules play an important role in stabilizing this antigen-antibody complex. The equilibrium constant for the binding of the mutant Fv to the antigen decreases by three orders of magnitude (from 2.3 x 10(8) M-1 to 2.6 x 10(5) M-1). Titration calorimetry shows that this results from a smaller negative binding enthalpy (delta delta H = -16 kJ mol-1 at 24 degrees C), whereas the value of the binding entropy is not affected. Since in the complex between the mutated Fv and antigen the buried area has decreased relative to that of the wild-type Fv by about 150 A2, the contribution of the buried unit area to the decrease in free energy (delta Gzero) is approximately 117 J mol-1 (28 cal mol-1) per A2. The loss of interatomic contacts in replacing Trp by Asp permits an approximate calculation for the contribution of van der Waals interactions made by Trp92 in this complex, which gives an average of 2.1 kJ mol-1 (0.5 kcal mol-1) for contacts between carbon atoms.
Publisher: Wiley
Date: 09-1995
Publisher: Elsevier BV
Date: 09-1994
Abstract: A complex between the Fv fragment of an anti-hen eggwhite lysozyme antibody (D1.3) and the Fv fragment of an antibody specific for an idiotypic determinant of D1.3 has been crystallized in a form suitable for X-ray diffraction analysis. Both Fv fragments were expressed in soluble form in Escherichia coli and purified by affinity chromatography diffraction-quality crystals were only obtained following separation of each Fv into distinct isoelectric forms. The crystals belong to space group C2, have unit cell dimensions a = 152.8 A, b = 79.4 A, c = 51.5 A, beta = 100.2 degrees, and diffract to better than 2.2 A resolution. The solvent content of the crystals is approximately 60% (v/v) with one Fv-Fv complex in the asymmetric unit. The ability to readily express both components of an antigen-antibody system in bacteria will allow us to rigorously assess the energetic contribution of in idual amino acids to complex formation through pairwise mutagenesis of interacting residues.
Publisher: Elsevier BV
Date: 03-2004
DOI: 10.1016/J.STR.2003.12.002
Abstract: Superantigens are a class of microbial proteins with the ability to excessively activate T cells by binding to the T cell receptor. The staphylococcal and streptococcal superantigens are closely related in structure and possess an N-terminal domain that resembles an OB fold and a C-terminal domain similar to a beta-grasp fold. Yersinia pseudotuberculosis produces superantigens, YPMa, YPMb, and YPMc, which have no significant amino acid similarity to other proteins. We have determined the crystal and solution structures of YPMa, which show that the protein has a jelly-roll fold. The closest structural neighbors to YPMa are viral capsid proteins and members of the tumor necrosis factor superfamily. In the crystal structure, YPMa packs as a trimer, another feature shared with viral capsid proteins and TNF superfamily proteins. However, in solution YPMa behaves as a monomer, and any functional relevance of the trimer observed in the crystals is yet to be established.
Publisher: Elsevier BV
Date: 03-1995
DOI: 10.1016/0031-6865(94)00046-X
Abstract: Antibodies possess a highly complementary combining site structure to that of their specific antigens. In many instances their reactions are driven by enthalpic factors including, at least in the case of the reaction of monoclonal antibody D1.3 with lysozyme, enthalpy of solvation. They require minor structural rearrangements, and their equilibrium association constants are relatively high (10(7)-10(11) M-1). By contrast, in an idiotope--anti-idiotope (antibody-antibody) reaction, which is entropically driven, the binding equilibrium constant is only 1.5 x 10(5) M-1 at 20 degrees C. This low value results from a slow association rate (10(3) M-1 s-1) due to a selection of conformational states that allow one of the interacting molecular surfaces (the idiotope on antibody D1.3) to become complementary to that of the anti-idiotopic antibody. Thus, antibody D1.3 reacts with two different macromolecules: with its specific antigen, hen egg lysozyme, and with a specific anti-idiotopic antibody. Complementarity with lysozyme is closer to a "lock and key" model and results in high affinity (2-4 x 10(8) M-1). That with the anti-idiotopic antibody involves conformational changes at its combining site and it results in a lower association constant (1.5 x 10(5) M-1). Thus, an "induced fit" mechanism may lead to a broadening of the binding specificity but with a resulting decrease in the intrinsic binding affinity which may weaken the physiological function of antibodies.
Publisher: Elsevier BV
Date: 07-1996
DOI: 10.1016/0167-5699(96)10020-7
Abstract: In recent years noninvasive brain stimulation (NIBS) applications have emerged as a third and novel treatment option alongside psychopharmacology and psychotherapy in the treatment of mental diseases. It is assumed that NIBS could represent a supplement or (in some indications) even replacement to established therapeutic strategies, e.g. in disorders with high resistance to current treatment regimens, such as negative symptoms or cognitive impairments in schizophrenia. Although positive symptoms in schizophrenia can be treated sufficiently with antipsychotic drugs, patients with negative symptoms frequently suffer from persistent lack of impetus, cognitive decline, social withdrawal and loss of global functioning in the activities of daily life however, in these cases, current treatment strategies exert only moderate effects, and new treatment options are urgently needed. This review article provides a summary of the clinical effects of new electrical NIBS methods, e.g. transcranial direct current stimulation (tDCS), transcranial alternating current stimulation (tACS), and transcranial random noise stimulation (tRNS) for the treatment of negative symptoms in schizophrenia. These new NIBS methods could help restore the disrupted neuronal networks and improve disturbed connectivity, especially of the left dorsolateral prefrontal cortex and left temporoparietal junction. Promising results are reported for the treatment of negative symptoms with tDCS, tACS and tRNS and could thus represent new therapeutic options in the treatment of schizophrenia.
Publisher: International Union of Crystallography (IUCr)
Date: 09-1994
Publisher: Springer Science and Business Media LLC
Date: 11-1996
DOI: 10.1038/384188A0
Abstract: Superantigens (SAgs) are viral or bacterial proteins that act as potent T-cell stimulants and have been implicated in a number of human diseases, including toxic shock syndrome, diabetes mellitus and multiple sclerosis. The interaction of SAgs with the T-cell receptor (TCR) and major histocompatibility complex (MHC) proteins results in the stimulation of a disproportionately large fraction of the T-cell population. We report here the crystal structures of the beta-chain of a TCR complexed with the Staphylococcus aureus enterotoxins C2 and C3 (SEC2, SEC3). These enterotoxins, which cause both toxic shock and food poisoning, bind in an identical way to the TCR beta-chain. The complementarity-determining region 2 (CDR2) of the beta-chain and, to lesser extents, CDR1 and hypervariable region 4 (HV4), bind in a cleft between the two domains of the SAgs. Thus, there is considerable overlap between the SAg-binding site and the peptide/MHC-binding sites of the TCR. A model of a TCR-SAg-MHC complex constructed from the crystal structures of (1) the beta-chain-SEC3 complex, (2) a complex between staphylococcal enterotoxin B (SEB) and an MHC molecule, and (3) a TCR V(alpha) domain, reveals that the SAg acts as a wedge between the TCR and MHC to displace the antigenic peptide away from the TCR combining site. In this way, the SAg is able to circumvent the normal mechanism for T-cell activation by specific peptide/MHC complexes.
Publisher: Elsevier BV
Date: 04-1996
Abstract: The crystal structure of the Fv fragment of the murine monoclonal anti-lysozyme antibody D1.3, complexed with turkey egg-white lysozyme (TEL), is presented. D1.3 (IgG1, kappa) is a secondary response antibody specific for hen egg-white lysozyme (HEL). TEL and HEL are homologous and differ in amino acid sequence in the antibody-antigen interface only at position 121. The side-chain of HEL residue Gln121 makes a pair of hydrogen bonds to main-chain atoms of the antibody light chain. In the D1.3-TEL structure, TEL residue His121 makes only one hydrogen bond with the light chain as a result of 129 degree and 145 degree change in peptide torsion angles for residues Trp92 and Ser93. Probably as a consequence of this conformational change, the D1.3-TEL association occurs at a much slower rate than the D1.3-HEL association. The D1.3-TEL complex is destabilized with respect to the D1.3-HEL interaction by the loss of two hydrogen bonds, exclusively due to the substitution of histidine for glutamine. While antibodies of secondary responses are indeed highly specific for antigen, this work demonstrates that by undergoing subtle conformational change antibodies can still recognize mutated protein antigens, albeit at a cost to affinity.
Publisher: Wiley
Date: 12-1996
Abstract: T-lymphocytes recognize a wide variety of antigens through highly erse cell-surface glycoproteins known as T-cell receptors (TCRs). These disulfide-linked heterodimers are composed of alpha and beta or gamma and delta polypeptide chains consisting of variable (V) and constant (C) domains non-covalently associated with at least four invariant chains to form the TCR-CD3 complex. It is well established that alpha beta TCRs recognize antigen in the form of peptides bound to molecules of the major histocompatibility complex (MHC) furthermore, information on the three-dimensional structure of alpha beta TCRs has recently become available through X-ray crystallography. In contrast, the antigen specificity of gamma delta TCRs is much less well understood and their three-dimensional structure is unknown. We have cloned the delta chain of a human TCR specific for the MHC class I HLA-A2 molecule and expressed the V domain as a secreted protein in the periplasmic space of Escherichia coli. Following affinity purification using a nickel chelate adsorbent, the recombinant V delta domain was crystallized in a form suitable for X-ray diffraction analysis. The crystals are orthorhombic, space group P2(1)2(1)2 with unit cell dimensions a = 69.9, b = 49.0, c = 61.6 A. and diffract to beyond 2.3 A resolution. The ability of a V delta domain produced in bacteria to form well-ordered crystals strongly suggests that the periplasmic space can provide a suitable environment for the correct in vivo folding of gamma delta TCRs.
Publisher: Elsevier BV
Date: 11-1996
Abstract: Anti-idiotopic antibodies react with unique antigenic features, usually associated with the combining sites, of other antibodies. They may thus mimic specific antigens that react with the same antibodies. The structural basis of this mimicry is analyzed here in detail for an anti-idiotopic antibody that mimics the antigen, hen egg-white lysozyme. The crystal structure of an anti-hen-egg-white lysozyme antibody (D1.3) complexed with an anti-idiotopic antibody (E5.2) has been determined at a nominal resolution of 1.9 A. E5.2 contacts substantially the same residues of D1.3 as lysozyme, thus mimicking its binding to D1.3. The mimicry embodies conservation of hydrogen bonding: six of the 14 protein-protein hydrogen bonds bridging D1.3-E5.2 are structurally equivalent to hydrogen bonds bridging D1.3-lysozyme. The mimicry includes a similar number of van der Waals interactions. The mimicry of E5.2 for lysozyme, however, does not extend to the topology of the non-polar surfaces of E5.2 and lysozyme, which are in contact with D1.3 as revealed by a quantitative analysis of the contacting surface similarities between E5.2 and lysozyme. The structure discussed herein shows that an anti-idiotopic antibody can provide an approximate topological and binding-group mimicry of an external antigen, especially in the case of the hydrophilic surfaces, even though there is no sequence homology between the anti-idiotope and the antigen.
Publisher: Wiley
Date: 03-1994
Abstract: The reaction between the mouse (BALB/c) anti-idiotopic monoclonal antibodies E225 and E5.2 and idiotopes on the (BALB/c) anti-lysozyme monoclonal antibody D1.3 has been characterized by titration calorimetry, by equilibrium sedimentation and by the determination of binding association and dissociation rates. The reaction between E5.2 and D1.3 is driven by a large negative enthalpy and its rate and equilibrium association constants are comparable to those observed in other antigen-antibody reactions. In contrast, the reaction between E225 and D1.3 is entropically driven and characterized by slow association kinetics (1 x 10(3) M-1 sec-1) and a resulting low equilibrium constant (Ka = 2 x 10(5) M-1). A correlation of these properties with the three-dimensional structure of the Fab225-FabD1.3 complex, previously determined by X-ray diffraction methods to 2.5 A resolution, indicates that conformational changes of several D1.3 contacting residues, located in its complementarity determining regions, may explain these features of the reaction.
Publisher: Springer Science and Business Media LLC
Date: 1998
DOI: 10.1038/35172
Abstract: Antigen recognition by T lymphocytes is mediated by cell-surface glycoproteins known as T-cell antigen receptors (TCRs). These are composed of alpha and beta, or gamma and delta, polypeptide chains with variable (V) and constant (C) regions. In contrast to alphabeta TCRs, which recognize antigen only as peptide fragments bound to molecules of the major histocompatibility complex (MHC), gammadelta TCRs appear to recognize proteins directly, without antigen processing, and to recognize MHC molecules independently of the bound peptide. Moreover, small phosphate-containing non-peptide compounds have also been identified as ligands for certain gammadelta T cells. These studies indicate that antigen recognition by gammadelta TCRs may be fundamentally different from that by alphabeta TCRs. The three-dimensional structures of several alphabeta TCRs and TCR fragments, and their complexes with peptide-MHC or superantigens, have been determined. Here we report the crystal structure of the Vdelta domain of a human gammadelta TCR at 1.9 A resolution. A comparison with antibody and alphabeta TCR V domains reveals that the framework structure of Vdelta more closely resembles that of VH than of Valpha, Vbeta or VL (where H and L refer to heavy and light chains), whereas the relative positions and conformations of its complementarity-determining regions (CDRs) share features of both Valpha and VH. These results provide the first direct evidence that gammadelta TCRs are structurally distinct from alphabeta TCRs and, together with the observation that the CDR3 length distribution of TCR delta chains is similar to that of immunoglobulin heavy chains, are consistent with functional studies suggesting that recognition of certain antigens by gammadelta TCRs may resemble antigen recognition by antibodies.
Publisher: Rockefeller University Press
Date: 12-1995
Abstract: The three-dimensional structure of an unglycosylated T cell antigen receptor (TCR) beta chain has recently been determined to 1.7 A resolution. To investigate whether this soluble beta chain (murine V beta 8.2J beta 2.1C beta 1) retains superantigen (SAG)-binding activity, we measured its affinity for various bacterial SAGs in the absence of MHC class II molecules. Dissociation constants (KDs) were determined using two independent techniques: surface plasmon resonance detection and sedimentation equilibrium. Specific binding was demonstrated to staphylococcal enterotoxins (SEs) B, C1, C2, and C3 and to streptococcal pyrogenic exotoxin A (SPEA), consistent with the known proliferative effects of these SAGs on T cells expressing V beta 8.2. In contrast, SEA, which does not stimulate V beta 8.2-bearing cells, does not bind the recombinant beta chain. Binding of the beta chain to SAGs was characterized by extremely fast dissociation rates (& 0.1 s-1), similar to those reported for certain leukocyte adhesion molecules. Whereas the beta chain bound SEC1, 2, and 3 with KDs of 0.9-2.5 microM, the corresponding value for SEB was approximately 140 microM. The much weaker binding to SEB than to SEC1, 2, or 3 was surprising, especially since SEB was found to actually be 3- to 10-fold more effective, on a molar basis, than the other toxins in stimulating the parental T cell hybridoma. We interpret these results in terms of the ability of SEC to activate T cells independently of MHC, in contrast to SEB. We have also measured SE binding to the glycosylated form of the beta chain and found that carbohydrate apparently does not contribute to recognition, even though the N-linked glycosylation sites at V beta 8.2 residues Asn24 and Asn74 are at or near the putative SAG-binding site. This result, along with the structural basis for the V beta specificity of SEs, are discussed in relation to the crystal structure of the unglycosylated beta chain.
No related grants have been discovered for Barry Fields.