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Toyoyuki Ose

Publications and source records attributed to Toyoyuki Ose.

12 recordsLinked to original sources
PubMed2006

Crystal structure of the human monocyte-activating receptor, "Group 2" leukocyte Ig-like receptor A5 (LILRA5/LIR9/ILT11).

Human leukocyte Ig-like receptor B1 (LILRB1) and B2 (LILRB2) belong to "Group 1" receptors and recognize a broad range of major histocompatibility complex class I molecules (MHCIs). In contrast, "Group 2" receptors show low similarity with LILRB1/B2, and their ligands remain to be identified. To date, the structural and functional characteristics of Group 2 LILRs are poorly understood. Here we report the crystal structure of the extracellular domain of LILRA5, which is an activating Group 2 LILR expressed on monocytes and neutrophils. Unexpectedly, the structure showed large changes in structural conformation and charge distribution in the region corresponding to the MHCI binding site of LILRB1/B2, which are also distinct from killer cell Ig-like receptors and Fc alpha receptors. These changes probably confer the structural hindrance for the MHCI binding, and their key amino acid substitutions are well conserved in Group 2 LILRs. Consistently, the surface plasmon resonance and flow cytometric analyses demonstrated that LILRA5 exhibited no affinities to all tested MHCIs. These results raised the possibility that LILRA5 as well as Group 2 LILRs do not play a role in any MHCI recognition but could possibly bind to non-MHCI ligand(s) on the target cells to provide a novel immune regulation mechanism.

Amino Acid Sequence↗
PubMed2006

Efficient leukocyte Ig-like receptor signaling and crystal structure of disulfide-linked HLA-G dimer.

HLA-G is a nonclassical major histocompatibility complex class I (MHCI) molecule, which is expressed in trophoblasts and confers immunological tolerance in the maternal-fetal interface by binding to leukocyte Ig-like receptors (LILRs, also called as LIR/ILT/CD85) and CD8. HLA-G is expressed in disulfide-linked dimer form both in solution and at the cell surface. Interestingly, MHCI dimer formations have been involved in pathogenesis and T cell activation. The structure and receptor binding characteristics of MHCI dimers have never been evaluated. Here we performed binding studies showing that the HLA-G dimer exhibited higher overall affinity to LILRB1/2 than the monomer by significant avidity effects. Furthermore, the cell reporter assay demonstrated that the dimer formation remarkably enhanced the LILRB1-mediated signaling at the cellular level. We further determined the crystal structure of the wild-type dimer of HLA-G with the intermolecular Cys(42)-Cys(42) disulfide bond. This dimer structure showed the oblique configuration to expose two LILR/CD8-binding sites upward from the membrane easily accessible for receptors, providing plausible 1:2 (HLA-G dimer:receptors) complex models. These results indicated that the HLA-G dimer conferred increased avidity in a proper structural orientation to induce efficient LILR signaling, resulting in the dominant immunosuppressive effects. Moreover, structural and functional implications for other MHCI dimers observed in activated T cells and the pathogenic allele, HLA-B27, are discussed.

Alleles↗
PubMed2005

Crystallization and preliminary crystallographic analysis of the N-terminal domain of PriA from Escherichia coli.

PriA, a DEXH-type DNA helicase, binds specifically to the 3' end of DNA through its N-terminal domain, and is a candidate sensor protein that recognizes arrested DNA replication forks in bacteria. We crystallized an N-terminal fragment of PriA in the absence and the presence of oligonucleotides to elucidate the structural basis for the specific recognition of the 3' terminus of DNA.

Adenosine Triphosphatases↗
PubMed2005

Crystallization and preliminary X-ray analysis of mitochondrial presequence receptor Tom20 in complexes with a presequence from aldehyde dehydrogenase.

Most mitochondrial proteins are synthesized in the cytosol and must be imported into the mitochondria. Many mitochondrial precursor proteins have an extra leader sequence at their N-terminus called a presequence. Presequences are recognized by the Tom20 receptor protein. Based on the previously determined NMR structure of rat Tom20, a fragment corresponding to the core structure was generated. A cysteine residue was added at the C-terminus of the rat aldehyde dehydrogenase presequence to fix the presequence peptide onto the Tom20 fragment via an intermolecular disulfide bond. Two crystal forms of the complex were successfully obtained with different designs of the linker sequence which diffracted to 2.1 and 1.9 A. Crystal dehydration and subsequent annealing was essential to obtain good diffraction data for the 2.1 A crystal form.

Aldehyde Dehydrogenase↗
PubMed2005

Crystallization and preliminary X-ray analysis of the mRNA-binding domain of elongation factor SelB in complex with RNA.

In bacteria, the selenocysteine-specific elongation factor SelB is necessary for incorporation of selenocysteine, the 21st amino acid, into proteins by the ribosome. SelB binds to an mRNA hairpin formed by the selenocysteine-insertion sequence (SECIS) and delivers selenocysteyl-tRNA (Sec-tRNASec) at the ribosomal A site. The minimum fragment (residues 512-634) of Moorella thermoacetica SelB (SelB-M) required for mRNA binding has been overexpressed and purified. The complex of SelB-M with 23 nucleotides of the SECIS mRNA hairpin was crystallized at 293 K using the hanging-drop vapour-diffusion or oil-batch methods. The crystals diffract to 2.3 A resolution using SPring-8 BL41XU and belong to the space group P2(1)2(1)2, with unit-cell parameters a = 81.69, b = 169.58, c = 71.69 A.

Bacterial Proteins↗
PubMed2005

Crystal structure of a biologically functional form of PriB from Escherichia coli reveals a potential single-stranded DNA-binding site.

PriB is not only an essential protein necessary for the replication restart on the collapsed and disintegrated replication fork, but also an important protein for assembling of primosome onto PhiX174 genomic DNA during replication initiation. Here we report a 2.0-A-resolution X-ray structure of a biologically functional form of PriB from Escherichia coli. The crystal structure revealed that despite a low level of primary sequence identity, the PriB monomer, as well as the dimeric form, are structurally identical to the N-terminal DNA-binding domain of the single-stranded DNA-binding protein (SSB) from Escherichia coli, which possesses an oligonucleotides-binding-fold. The oligonucleotide-PriB complex model based on the oligonucleotides-SSB complex structure suggested that PriB had a DNA-binding pocket conserved in SSB from Escherichia coli and might bind to single-stranded DNA in the manner of SSB. Furthermore, surface plasmon resonance analysis and fluorescence measurements demonstrated that PriB binds single-stranded DNA with high affinity, by involving tryptophan residue. The significance of these results with respect to the functional role of PriB in the assembly of primosome is discussed.

Amino Acid Sequence↗
PubMed2005

Structural basis for mRNA recognition by elongation factor SelB.

In bacteria, incorporation of selenocysteine, the 21(st) amino acid, into proteins requires elongation factor SelB, which has the unusual property of binding to both transfer RNA (tRNA) and mRNA. SelB binds to an mRNA hairpin formed by the selenocysteine insertion sequence (SECIS) with extremely high specificity, the molecular basis of which has been unknown. We have determined the crystal structure of the mRNA-binding domain of SelB in complex with SECIS RNA at a resolution of 2.3 A. This is the first example of a complex between an RNA and a winged-helix (WH) domain, a motif found in many DNA-binding proteins and recently discovered in RNA-binding proteins. Notably, RNA binding does not induce a major conformational change in the WH motif. The structure reveals a new mode of RNA recognition with a geometry that allows the complex to wrap around the small ribosomal subunit.

Amino Acid Motifs↗
PubMed2005

Crystallization and preliminary X-ray analysis of alpha-xylosidase from Escherichia coli.

Glycoside hydrolases have been implicated in many biological processes. To date, they have been classified into 93 glycoside hydrolase (GH) families based on amino-acid sequence similarity. alpha-Xylosidase from Escherichia coli belongs to GH family 31 and catalyzes the release of alpha-xylose from the non-reducing terminal side of alpha-xyloside. Single crystals of alpha-xylosidase have been grown by vapour diffusion at 293 K from 10%(w/v) PEG 20K, 2%(v/v) 2-propanol, 2%(v/v) glycerol and 0.1 M 2-morpholinoethanesulfonic acid pH 5.5. These crystals belong to space group P2(1)2(1)2(1) and X-ray diffraction data were collected to a resolution of 2.75 A. Crystals of selenomethionyl-substituted alpha-xylosidase were also obtained, which diffracted to at least 3.0 A. Based on the value of VM, the asymmetric unit in these crystals was assumed to contain six molecules.

Escherichia coli↗
PubMed2004

Structure of macrophomate synthase.

Macrophomate synthase (MPS) is an enzyme that catalyzes an extraordinarily complex conversion reaction, including two decarboxylations, two carbon-carbon bond formations and a dehydration, to form the benzoate analogue macrophomate from a 2-pyrone derivative and oxalacetate. Of these reactions, the two carbon-carbon bond formations are especially noteworthy because previous experiments have indicated that they proceed via a Diels-Alder reaction, one of the most widely used reactions in organic synthesis. The structural evidence that MPS catalyzes an intermolecular Diels-Alder reaction has been reported recently [Ose et al. (2003), Nature (London), 422, 185-189]. Interestingly, the tertiary structure as well as the quaternary structure of MPS are similar to those of 2-dehydro-3-deoxygalactarate (DDG) aldolase, a carbon-carbon bond-forming enzyme that catalyzes the reversible reaction of aldol condensation/cleavage. Here, the structure of MPS is described in detail and compared with that of DDG aldolase. Both enzymes have a (beta/alpha)(8)-barrel fold and are classified as belonging to the enolase superfamily based on their reaction strategy. The basic principles for carbon-carbon bond formation used by both MPS and DDG aldolase are the same with regard to trapping the enolate substrate and inducing subsequent reaction. The major differences in the active sites between these two enzymes are the recognition mechanisms of the second substrates, 2-pyrone and DDG, respectively.

Aldehyde-Lyases↗
PubMed2003

Reaction intermediate structures of 1-aminocyclopropane-1-carboxylate deaminase: insight into PLP-dependent cyclopropane ring-opening reaction.

The pyridoxal 5'-phosphate-dependent enzymes have been evolved to catalyze diverse substrates and to cause the reaction to vary. 1-Aminocyclopropane-1-carboxylate deaminase catalyzes the cyclopropane ring-opening reaction followed by deamination specifically. Since it was discovered in 1978, the enzyme has been widely investigated from the mechanistic and physiological viewpoints because the substrate is a precursor of the plant hormone ethylene and the enzymatic reaction includes a cyclopropane ring-opening. We have previously reported the crystal structure of the native enzyme. Here we report the crystal structures of the two reaction intermediates created by the mutagenesis complexed with the substrate. The substrate was validated in the active site of two forms: 1). covalent-bonded external aldimine with the coenzyme in the K51T form and 2). the non-covalent interaction around the coenzyme in the Y295F form. The orientations of the substrate in both structures were quite different form each other. In concert with other site-specific mutation experiments, this experiment revealed the ingenious and unique strategies that are used to achieve the specific activity. The substrate incorporated into the active site is reactivated by a two-phenol charge relay system to lead to the formation of a Schiff base with the coenzyme. The catalytic Lys51 residue may play a novel role to abstract the methylene proton from the substrate in cooperation with other factors, the carboxylate group of the substrate and the electron-adjusting apparatuses of the coenzyme.

Binding Sites↗
PubMed2003

Insight into a natural Diels-Alder reaction from the structure of macrophomate synthase.

The Diels-Alder reaction, which forms a six-membered ring from an alkene (dienophile) and a 1,3-diene, is synthetically very useful for construction of cyclic products with high regio- and stereoselectivity under mild conditions. It has been applied to the synthesis of complex pharmaceutical and biologically active compounds. Although evidence on natural Diels-Alderases has been accumulated in the biosynthesis of secondary metabolites, there has been no report on the structural details of the natural Diels-Alderases. The function and catalytic mechanism of the natural Diels-Alderase are of great interest owing to the diversity of molecular skeletons in natural Diels-Alder adducts. Here we present the 1.70 A resolution crystal structure of the natural Diels-Alderase, fungal macrophomate synthase (MPS), in complex with pyruvate. The active site of the enzyme is large and hydrophobic, contributing amino acid residues that can hydrogen-bond to the substrate 2-pyrone. These data provide information on the catalytic mechanism of MPS, and suggest that the reaction proceeds via a large-scale structural reorganization of the product.

Alcohols↗