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I A Wilson

Publications and source records attributed to I A Wilson.

At least 37 records · Page 2Linked to original sources
PubMed2001

Crystal structure of the murine NK cell-activating receptor NKG2D at 1.95 A.

NKG2D, a homodimeric lectin-like receptor, is a unique stimulatory molecule that is found on natural killer cells,T cells and activated macrophages. The natural ligands for murine NKG2D are distant major histocompatibility complex homologs, retinoic acid early transcript (Rae1) and H-60 minor histocompatibility antigen. The crystal structure of the extracellular region of murine NKG2D reveals close homology with other C-type lectin receptors such as CD94, Ly49A, rat MBP-A and CD69. However, the precise mode of dimeric assembly varies among these natural killer receptors, as well as their surface topography and electrostatic properties. The NKG2D structure provides the first structural insights into the role and ligand specificity of this stimulatory receptor in the innate and adaptive immune system.

Amino Acid Sequence↗
PubMed2001

Crystal structure of a bifunctional transformylase and cyclohydrolase enzyme in purine biosynthesis.

ATIC, the product of the purH gene, is a 64 kDa bifunctional enzyme that possesses the final two activities in de novo purine biosynthesis, AICAR transformylase and IMP cyclohydrolase. The crystal structure of avian ATIC has been determined to 1.75 A resolution by the MAD method using a Se-methionine modified enzyme. ATIC forms an intertwined dimer with an extensive interface of approximately 5,000 A(2) per monomer. Each monomer is composed of two novel, separate functional domains. The N-terminal domain (up to residue 199) is responsible for the IMPCH activity, whereas the AICAR Tfase activity resides in the C-terminal domain (200-593). The active sites of the IMPCH and AICAR Tfase domains are approximately 50 A apart, with no structural evidence of a tunnel connecting the two active sites. The crystal structure of ATIC provides a framework to probe both catalytic mechanisms and to design specific inhibitors for use in cancer chemotherapy and inflammation.

Animals↗
PubMed2001

Unraveling the mysteries of gammadelta T cell recognition.

gammadelta T cells are the misunderstood siblings of the antigen receptor family. A recent paper in Nature that describes the crystal structure of a gammadelta TCR should initiate a clearer understanding of these enigmatic cells.

Animals↗
PubMed2001

Crystallization and preliminary structure determination of an intact human immunoglobulin, b12: an antibody that broadly neutralizes primary isolates of HIV-1.

An intact human immunoglobulin with a full-length hinge has been crystallized for the first time in a form in which all of the Ig domains are ordered. The IgG1 antibody b12 is one of only three known monoclonal antibodies described that potently neutralize a broad range of HIV-1 primary isolates. It binds to an epitope overlapping the conserved CD4 binding site on the viral surface antigen gp120. Hexagonal crystals corresponding to space group R32 were grown from 0.8 M ammonium sulfate, with unit-cell parameters a = b = 271.3, c = 175.2 A and one molecule per asymmetric unit. The crystals diffract to 2.8 A and a preliminary molecular-replacement solution indicates that all 12 Ig domains of the antibody can be resolved.

Crystallization↗
PubMed2001

Preliminary X-ray diffraction studies of the transcriptional inhibitory antibody Fab41.4.

The binding of transcription factor ATF-1 to DNA contributes to gene expression and regulation of cell growth. Antibody Mab41.4, raised against ATF-1, and its derivatives Fab41.4 and scFv41.4 inhibit specific DNA binding in vitro and induce apoptotic death of tumor cells in vivo. Structural studies of Fab41.4 were performed to gain insight into the mechanism of action of this potentially therapeutic antibody. The optimal conditions for crystallization of Fab41.4 were determined. Crystals were needle-like in appearance, displayed C2 space-group symmetry and diffracted to a resolution of 1.6 A. The unit-cell parameters were determined to be a = 186.64, b = 40.22, c = 55.58 A, alpha = gamma = 90, beta = 96.93 degrees. The data set was 97.7% complete. Molecular replacement was performed, resulting in an R value of 44.6%.

Activating Transcription Factor 1↗
PubMed2000

Catalysis of decarboxylation by a preorganized heterogeneous microenvironment: crystal structures of abzyme 21D8.

Antibody 21D8 catalyzes the solvent-sensitive decarboxylation of 3-carboxybenzisoxazoles. The crystal structure of chimeric Fab 21D8 with and without hapten at 1.61 A and 2.10 A, respectively, together with computational analysis, shows how a melange of polar and non-polar sites are exploited to achieve both substrate binding and acceleration of a reaction normally facilitated by purely aprotic dipolar media. The striking similarity of the decarboxylase and a series of unrelated esterase antibodies also highlights the chemical versatility of structurally conserved anion binding sites and the relatively subtle changes involved in fine-tuning the immunoglobulin pocket for recognition of different ligands and catalysis of different reactions.

Amino Acid Sequence↗
PubMed2000

The I-Ag7 MHC class II molecule linked to murine diabetes is a promiscuous peptide binder.

Susceptibility to insulin-dependent diabetes mellitus is linked to MHC class II genes. The only MHC class II molecule expressed by nonobese diabetic (NOD) mice, I-Ag7, shares a common alpha-chain with I-Ad but has a peculiar beta-chain. As with most beta-chain alleles linked to diabetes susceptibility, I-Ag7 contains a nonaspartic residue at position beta57. We have produced large amounts of empty I-Ag7 molecules using a fly expression system to characterize its biochemical properties and peptide binding by phage-displayed peptide libraries. The identification of a specific binding peptide derived from glutamic acid decarboxylase (GAD65) has allowed us to crystallize and obtain the three-dimensional structure of I-Ag7. Structural information was critical in evaluating the binding studies. I-Ag7, like I-Ad, appears to be very promiscuous in terms of peptide binding. Their binding motifs are degenerate and contain small and/or small hydrophobic residues at P4 and P6 of the peptide, a motif frequently found in most globular proteins. The degree of promiscuity is increased for I-Ag7 over I-Ad as a consequence of a larger P9 pocket that can specifically accommodate negatively charged residues, as well as possibly residues with bulky side chains. So, although I-Ad and I-Ag7 are structurally closely related, stable molecules and good peptide binders, they differ functionally in their ability to bind significantly different peptide repertoires that are heavily influenced by the presence or the absence of a negatively charged residue at position 57 of the beta-chain. These characteristics link I-Ag7 with autoimmune diseases, such as insulin-dependent diabetes mellitus.

Amino Acid Motifs↗
PubMed2000

Structural requirements for antigen presentation by mouse CD1.

The structural basis for the T cell response to glycolipid antigens (Ags) remains poorly understood. T lymphocytes autoreactive for mouse CD1 (mCD1.1) or reactive for the glycosphingolipid alphagalactosylceramide (alpha-GalCer) presented by mCD1.1 have been described previously. In this paper it is shown that mutations at the top of the alpha helices and in the bottom of the Ag-binding groove can disrupt both mCD1.1 autoreactivity and alpha-GalCer recognition. The locations of the positions that affect T cell responses indicate that recognition of mCD1.1 is not likely to be unconventional or superantigen-like. Furthermore, the effects of the bottom of the pocket mutation suggest that the autoreactive response could require an autologous ligand, and they indicate that alpha-GalCer binds to the groove of mCD1.1, most likely with the shorter 18-carbon hydrophobic chain in the A' pocket. Natural killer T cell hybridomas with identical T cell antigen receptor (TCR) alpha chains and different beta chains respond differently to alpha-GalCer presented by mCD1.1 mutants. This finding indicates a role for TCR beta in defining natural killer T cell specificity, despite the more restricted diversity of the alpha chains in these cells. Overall, the data are consistent with a mode of lipoglycan recognition similar to that proposed for glycopeptides, in which the TCR alpha and beta chains survey a surface composed of both mCD1.1 and the carbohydrate portion of alpha-GalCer.

Amino Acid Substitution↗
PubMed2000

A structural framework for deciphering the link between I-Ag7 and autoimmune diabetes.

Susceptibility to murine and human insulin-dependent diabetes mellitus correlates strongly with major histocompatibility complex (MHC) class II I-A or HLA-DQ alleles that lack an aspartic acid at position beta57. I-Ag7 lacks this aspartate and is the only class II allele expressed by the nonobese diabetic mouse. The crystal structure of I-Ag7 was determined at 2.6 angstrom resolution as a complex with a high-affinity peptide from the autoantigen glutamic acid decarboxylase (GAD) 65. I-Ag7 has a substantially wider peptide-binding groove around beta57, which accounts for distinct peptide preferences compared with other MHC class II alleles. Loss of Asp(beta57) leads to an oxyanion hole in I-Ag7 that can be filled by peptide carboxyl residues or, perhaps, through interaction with the T cell receptor.

Alleles↗
PubMed2000

Using antibody catalysis to study the outcome of multiple evolutionary trials of a chemical task.

Catalytic aldolase antibodies generated by immunization with two different, but structurally related, beta-diketone haptens were cloned and sequenced to study similarities and differences between independently evolved catalysts. Kinetic and sequence analysis coupled with mutagenesis, structural, and modeling studies reveal that the defining event in the evolution of these catalysts was a somatic mutation that placed a lysine residue in a deep, yet otherwise unrefined, hydrophobic pocket. We suggest that covalent chemistries may be as readily selected from the immune repertoire as the traditional noncovalent interactions that have formed the basis of immunochemistry until this time. Further, we believe that these experiments recapitulate the defining events in the evolution of nature's enzymes, particularly as they relate to chemical mechanism, catalytic promiscuity, and gene duplication.

Amino Acid Sequence↗
PubMed2000

Crystal structure of a gammadelta T cell receptor ligand T22: a truncated MHC-like fold.

Murine T10 and T22 are highly related nonclassical major histocompatibility complex (MHC) class Ib proteins that bind to certain gammadelta T cell receptors (TCRs) in the absence of other components. The crystal structure of T22b at 3.1 angstroms reveals similarities to MHC class I molecules, but one side of the normal peptide-binding groove is severely truncated, which allows direct access to the beta-sheet floor. Potential gammadelta TCR-binding sites can be inferred from functional mapping of T10 and T22 point mutants and allelic variants. Thus, T22 represents an unusual variant of the MHC-like fold and indicates that gammadelta and alphabeta TCRs interact differently with their respective MHC ligands.

Alleles↗
PubMed2000

A functional hot spot for antigen recognition in a superagonist TCR/MHC complex.

A longstanding question in T cell receptor signaling is how structurally similar ligands, with similar affinities, can have substantially different biological activity. The crystal structure of the 2C TCR complex of H-2Kb with superagonist peptide SIYR at 2.8 A elucidates a structural basis for TCR discrimination of altered peptide ligands. The difference in antigen potency is modulated by two cavities in the TCR combining site, formed mainly by CDRs 3alpha, 3beta, and 1beta, that complement centrally located peptide residues. This "functional hot spot" allows the TCR to finely discriminate amongst energetically similar interactions within different ligands for those in which the peptide appropriately stabilizes the TCR/pMHC complex and provides a new structural perspective for understanding differential signaling resulting from T cell cross-reactivity.

Animals↗
PubMed2000

A new functional domain of guanine nucleotide dissociation inhibitor (alpha-GDI) involved in Rab recycling.

Guanine nucleotide dissociation inhibitor (GDI) is a 55-kDa protein that functions in vesicular membrane transport to recycle Rab GTPases. We have now determined the crystal structure of bovine alpha-GDI at ultra-high resolution (1.04 A). Refinement at this resolution highlighted a region with high mobility of its main-chain residues. This corresponded to a surface loop in the primarily alpha-helical domain II at the base of alpha-GDI containing the previously uncharacterized sequence-conserved region (SCR) 3A. Site-directed mutagenesis showed that this mobile loop plays a crucial role in binding of GDI to membranes and extraction of membrane-bound Rab. This domain, referred to as the mobile effector loop, in combination with Rab-binding residues found in the multi-sheet domain I at the apex of alpha-GDI may provide flexibility for recycling of diverse Rab GTPases. We propose that conserved residues in domains I and II synergize to form the functional face of GDI, and that domain II mediates a critical step in Rab recycling during vesicle fusion.

Amino Acid Motifs↗
PubMed2000

Crystallization and preliminary crystallographic investigations of avian 5-aminoimidazole-4-carboxamide ribonucleotide transformylase-inosine monophosphate cyclohydrolase expressed in Escherichia coli.

ATIC [5-aminoimidazole-4-carboxamide ribonucleotide transformylase (AICAR Tfase)-inosine monophosphate cyclohydrolase (IMPCH)] is a bifunctional enzyme that catalyzes the penultimate and final steps in the de novo purine biosynthesis pathway and thus is an attractive anticancer target. Recombinant avian ATIC has been purified from an Escherichia coli expression system and crystallized in a binary complex with methotrexate (MTX). Crystals were obtained from PEG 4000 or MPEG 5000 buffered at pH 7.0-7.2 and data were collected from a single crystal at 96 K to 2.3 A resolution at the Stanford Synchrotron Radiation Laboratory (SSRL). The crystals are monoclinic and belong to space group P2(1), with unit-cell dimensions a = 65.17, b = 105.93, c = 103.47 A, beta = 108.27 degrees. Assuming two molecules per asymmetric unit, the Matthews coefficient V(m) is 2.63 A(3) Da(-1) and the solvent volume is 52.9%.

Animals↗
PubMed2000

Structural implications for the design of molecular vaccines.

The major histocompatibility complex molecules bind and present short antigenic peptide fragments on the surface of antigen presenting cells to T-cell receptors. Recognition of peptide-MHC by cytotoxic T-cells initiates a cascade of signals to T-cells, which in turn destroy the antigen presenting cell. In the design of molecular vaccines for the treatment of diseases, an understanding of the 3-dimensional structure of MHC class I and is interaction with both peptide and T-cell receptor is an important prerequisite. In this review, we will discuss such crystal structures, as well as structures of glycopeptides and alternative T-cell antigens presented by MHC molecules.

Amino Acid Sequence↗
PubMed1999

New insights into inhibitor design from the crystal structure and NMR studies of Escherichia coli GAR transformylase in complex with beta-GAR and 10-formyl-5,8,10-trideazafolic acid.

The crystal structure of Escherichia coli GAR Tfase at 2.1 A resolution in complex with 10-formyl-5,8,10-trideazafolic acid (10-formyl-TDAF, K(i) = 260 nM), an inhibitor designed to form an enzyme-assembled multisubstrate adduct with the substrate, beta-GAR, was studied to determine the exact nature of its inhibitory properties. Rather than forming the expected covalent adduct, the folate inhibitor binds as the hydrated aldehyde (gem-diol) in the enzyme active site, in a manner that mimics the tetrahedral intermediate of the formyl transfer reaction. In this hydrated form, the inhibitor not only provides unexpected insights into the catalytic mechanism but also explains the 10-fold difference in inhibitor potency between 10-formyl-TDAF and the corresponding alcohol, and a further 10-fold difference for inhibitors that lack the alcohol. The presence of the hydrated aldehyde was confirmed in solution by (13)C-(1)H NMR spectroscopy of the ternary GAR Tfase-beta-GAR-10-formyl-TDAF complex using the (13)C-labeled 10-formyl-TDAF. This insight into the behavior of the inhibitor, which is analogous to protease or transaminase inhibitors, provides a novel and previously unrecognized basis for the design of more potent inhibitors of the folate-dependent formyl transfer enzymes of the purine biosynthetic pathway and development of anti-neoplastic agents.

Binding, Competitive↗
PubMed1999

Evolution of shape complementarity and catalytic efficiency from a primordial antibody template.

The crystal structure of an efficient Diels-Alder antibody catalyst at 1.9 angstrom resolution reveals almost perfect shape complementarity with its transition state analog. Comparison with highly related progesterone and Diels-Alderase antibodies that arose from the same primordial germ line template shows the relatively subtle mutational steps that were able to evolve both structural complementarity and catalytic efficiency.

Antibodies, Catalytic↗