Showing papers by "Yoshiki Yamaguchi published in 2010"
TL;DR: The present study shows that brain endothelial cells expressed APP770, a differently spliced APP mRNA isoform from neuronal APP695, and produced Aβ40 and Aβ42, and found that the endothelial APP770 had sialylated core 1 type O-glycans.
93 citations
TL;DR: Biochemical assays support a model in which the self-assembled UbcH5b~Ub can serve as a bridge for the gap between the lysine residue of the substrate and the catalytic cysteine of E2, enabling efficient ubiquitination of substrates.
88 citations
TL;DR: Crystal structure of a human OS-9 MRH domain (OS-9(MRH)) complexed with α3,α6-mannopentaose has a flattened β-barrel structure with a characteristic P-type lectin fold and possesses distinctive double tryptophan residues in the oligosaccharide-binding site.
85 citations
TL;DR: A mechanistic model of PDI action is proposed; the a' domain transfers its own disulfide bond into the unfolded protein accommodated on the hydrophobic surface of the substrate-binding region, which consequently changes into a "closed" form releasing the oxidized substrate.
60 citations
TL;DR: NMR data revealed that the sugar–lipid interface is primarily perturbed upon binding of Aβ to the micelles, underscoring the importance of the inner part of the ganglioside cluster for accommodating Aβ in comparison with the outer carbohydrate branches that provide microbial toxin‐ and virus‐binding sites.
57 citations
48 citations
TL;DR: It is reported that columnar molecular hosts with parallel-aligned aromatics are magnetically oriented and exhibit observable RDC, and, upon encapsulation, an RDC silent guest is placed in alignment with the host and magnetic field and displays detectable induced RDC.
Abstract: 3D structural information can be obtained from the residual dipolar coupling (RDC) from biomolecules trapped within alignment media (e.g., liquid crystals). Unfortunately, RDC is not a suitable method for small molecules, as the intermolecular interactions with common alignment media are weak and ineffective. We report that columnar molecular hosts with parallel-aligned aromatics are magnetically oriented and exhibit observable RDC. And, upon encapsulation, an RDC silent guest (pyrene) is placed in alignment with the host and magnetic field and displays detectable induced RDC. Thus, the discrete hosts with aligned aromatic panels behave as molecular alignment media or “magnetic aligners” for small molecules. This method will be further applied to the 3D structure determination of more complex guest molecules.
35 citations
TL;DR: In this paper, aromatic ketones are enantioseletively hydrogenated in alcohols containing [RuX{(S,S)-Tsdpen)(eta(6)-p-cymene)] as precatalysts.
Abstract: Aromatic ketones are enantioseletively hydrogenated in alcohols containing [RuX{(S,S)-Tsdpen)(eta(6)-p-cymene)] (Tsdpen=TsNCH(C6H5)CH-(C6H3)NH2; X = TfO, Cl) as precatalysts. The corresponding Ru hydride (X = H) acts as a reducing species. The solution structures and complete spectral assignment of these complexes have been determined using 2D NMR (H-1-H-1 DQF-COSY, H-1-C-13 HMQC, HSQC, and H-1-F-19 HOESY). Depending on the nature of the solvents and conditions, the precatalysts exist as a covalently bound complex, tight ion pair of [Ru+(Tsdpen)(cymene)] and X-, solvent-separated ion pair, or discrete free ions. Solvent effects on the NH2 chemical shifts of the Ru complexes and the hydrodynamic radius and volume of the Re and TfO- ions elucidate the process of precatalyst activation for hydrogenation. Most notably, the Ru triflate possessing a high ionizability, substantiated by cyclic voltammetry, exists in alcoholic solvents largely as a solvent-separated ion pair and/or free ions. Accordingly, its diffusion-derived data in CD3OD reflect the independent motion of [Ru+(Tsdpen)(cymene) and TfO-. In CDCl3, the complex largely retains the covalent structure showing similar diffusion data for the cation and anion. The Ru triflate and chloride show similar but distinct solution behavior in various solvents. Conductivity measurements and catalytic behavior demonstrate that both complexes ionize in CH3OH to generate a common [Ru+(Tsdpen)(cymene)] and X-, although the extent is significantly greater for X = TfO-. The activation of [RuX(Tsdpen)(cymene)] during catalytic hydrogenation in alcoholic solvent occurs by simple ionization to generate [Ru+(Tsdpen)(cymene)]. The catalytic activity is thus significantly influenced by the reaction conditions.
34 citations
TL;DR: It is suggested that the conformation of PDI is controlled by the redox state of the active site cysteine residues in the a and a' domains and that the conformational alternation accompanies solvation changes in the activeSite cleft formed by the a, b, b', and a', domains.
Abstract: Protein disulfide isomerase (PDI) acts as folding catalyst and molecular chaperone for disulfide-containing proteins through the formation, breakage, and rearrangement of disulfide bonds. PDI has a modular structure comprising four thioredoxin domains, a, b, b′, and a′, followed by a short segment, c. The a and a′ domains have an active site cysteine pair for the thiol−disulfide exchange reaction, which alters PDI between the reduced and oxidized forms, and the b′ domain provides a primary binding site for substrate proteins. Although the structures and functions of PDI have studied, it is still argued whether the overall conformation of PDI depends on the redox state of the active site cysteine pair. Here, we report redox-dependent conformational and solvation changes of PDI from a thermophilic fungus elucidated by small-angle X-ray scattering (SAXS) analysis. The redox state and secondary structures of PDI were also characterized by nuclear magnetic resonance and circular dichroic spectroscopy, respecti...
34 citations
TL;DR: Analysis of the solution structure of ARMET suggests that ARMET may hold a negatively charged molecule using the two positively charged domains.
32 citations
TL;DR: It is demonstrated that GnT-III transfection has the potential to be an effective approach in humanizing the N-glycosylation of lepidopteran insect cells, thereby providing a possible preliminary step for the generation of complex-type glycoforms if the presence of a bisecting GlcNAc can be tolerated.
Abstract: The baculovirus-insect cell expression system is in widespread use for expressing post-translationally modified proteins. As a result, it is potentially applicable for the production of glycoproteins for therapeutic and diagnostic purposes. For practical use, however, remodeling of the biosynthetic pathway of host-cell N-glycosylation is required because insect cells produce paucimannosidic glycoforms, which are different from the typical mammalian glycoform, due to trimming of the non-reducing terminal beta1,2-GlcNAc residue of the core structure by a specific beta-N-acetylglucosaminidase. In order to establish a cell line which could be used as a host for the baculovirus-based production of glycoproteins with mammalian-type N-glycosylation, we prepared and characterized Spodoptera frugiperda Sf21 cells that had been transfected with the rat cDNA for beta1,4-N-acetylglucosaminyltransferase III (GnT-III), which catalyzes the addition of a bisecting GlcNAc. As evidenced by structural analyses of N-glycans prepared from whole cells and the expressed recombinant glycoproteins, the introduction of GnT-III led to the production of bisected hybrid-type N-glycans in which the beta1,2-GlcNAc residue at the alpha1,3-mannosyl branch is completely retained and which has the potential to be present in mammalian cells. These results and other related findings suggest that bisected oligosaccharides are highly resistant to beta-N-acetylglucosaminidase activity of the S. frugiperda fused lobes gene product, or other related enzymes, which was confirmed in Sf21 cells. Our present study demonstrates that GnT-III transfection has the potential to be an effective approach in humanizing the N-glycosylation of lepidopteran insect cells, thereby providing a possible preliminary step for the generation of complex-type glycoforms if the presence of a bisecting GlcNAc can be tolerated.
TL;DR: It is proposed that the N-terminal region of alpha-synuclein plays a key role in the formation ofalpha-syn DNA assemblies, and ultra-high-field NMR analysis of inhibitor-bound alpha- Synuclein dimers showed that the signals derived from the N/A region exhibited line broadening, confirming that theN-terminals is involved in inhibitor-induced dimerization.
TL;DR: Stable-isotope-assisted NMR analyses of immunoglobulin G-Fc glycoproteins and the glycopeptides derived therefrom are presented in terms of the dynamics and interactions of glycoconjugates.
Abstract: Unique advantages offered by nuclear magnetic resonance (NMR) spectroscopy provide high-resolution information not only on structures but also on dynamics and interactions of glycoconjugates in solution. These benefits are further enhanced by applying stable-isotope-labeling techniques, which we have developed. Our stable-isotope-assisted NMR analyses of immunoglobulin G-Fc glycoproteins and the glycopeptides derived therefrom are here presented in terms of the dynamics and interactions of glycoconjugates.
TL;DR: Findings suggest that FUT8 catalyzes the fucosylation of chitooligosaccharide analogous to NodZ, but that a nonreducing terminal chitotriose structure is required for the reaction.
Abstract: FUT8, a eukaryotic alpha1,6-fucosyltransferase, catalyzes the transfer of a fucosyl residue from guanine nucleotide diphosphate-beta-l-fucose to the innermost GlcNAc of an asparagine-linked oligosaccharide (N-glycan). The catalytic domain of FUT8 is structurally similar to that of NodZ, a bacterial alpha1,6-fucosyltransferase, which acts on a chitooligosaccharide in the synthesis of Nod factor. While the substrate specificities for the nucleotide sugar and the N-glycan have been determined, it is not known whether FUT8 is able to fucosylate other sugar chains such as chitooligosaccharides. The present study was conducted to investigate the action of FUT8 on chitooligosaccharides that are not generally thought to be a substrate in mammals, and the results indicate that FUT8 is able to fucosylate such structures in a manner comparable to NodZ. Surprisingly, structural analyses of the fucosylated products by high performance liquid chromatography, mass spectrometry and nuclear magnetic resonance indicated that FUT8 does not utilize the reducing terminal GlcNAc for fucose transfer but shows a preference for the third GlcNAc residue from the nonreducing terminus of the acceptor. These findings suggest that FUT8 catalyzes the fucosylation of chitooligosaccharide analogous to NodZ, but that a nonreducing terminal chitotriose structure is required for the reaction. The substrate recognition by which FUT8 selects the position to fucosylate might be distinct from that for NodZ and could be due to structural factor requirements which are inherent in FUT8.
TL;DR: It is reported that clustering of Siglec-7 with a specific F(ab')(2) elicited cell death, and the signal for which is transduced by an extracellular C2-set domain.
Abstract: Siglec-7, a sialic acid binding immunoglobulin-like lectin, predominantly transduces inhibitory signals through cytosolic immunoreceptor tyrosine-based inhibitory motifs (ITIMs). Here, we report that clustering of Siglec-7 with a specific F(ab')(2) elicited cell death. Interestingly, a truncated Siglec-7 lacking the cytosolic ITIM domain still induced the cell death, suggesting that the ITIMs are not essential for the death signaling. Further analyses of the death signaling revealed that an oxygen radical scavenger, N-acetyl cysteine, completely inhibited the cell death, whereas a pancaspase inhibitor did not. In addition, caspase-3 activation, DNA ladder formation, and nuclear condensation were not detected during the death process, suggesting that the cell death is nonapoptotic. To identify the critical region for the death signaling, we prepared a series of shuffling chimeras between Siglec-7 and Siglec-9, the latter of which did not transduce a death signal. The critical region was mapped to the middle of the membrane-proximal C2-set domain, which contained only six amino acid differences between Siglec-7 and Siglec-9. Point mutation analyses of each of these six amino acids revealed that four of the six amino acids were critical for the death signal. A computer-assisted 3D modeling revealed that these four amino acids were proximally located on the surface of the C2-set domain. In conclusion, Siglec-7 induces nonapoptotic cell death, the signal for which is transduced by an extracellular C2-set domain.
TL;DR: NMR data suggest that functional interactions of OPN with integrins and the other binding partners can be modulated by the intramolecular interactions.
TL;DR: It is suggested that the flexible nature of the isopeptide linkage thus observed contributes to the structural arrangements of ubiquitin chains exemplified by the pH-dependent closed-to-open conformational transition of tetraubiquitin.
TL;DR: The binding epitope of AG2 pentasaccharide to human Siglec-2 is determined, as the Galpalpha(1-4)Neu5Acalpha(2-3)Galp unit.
TL;DR: It is found that a major part of this mutant is sulfated at Tyr296, which adjacently precedes the originally glycosylated site, demonstrating that mutational deglycosylation can induce an unexpected post‐translational modification in the protein.
01 Jan 2010
TL;DR: Eri Sakata, Tadashi SATOH, Shunsuke YAMAMOTO, Yoshiki YAMAGUCHI, Maho YAGI-UTSUMI, Eiji KURIMOTO, Keiji TANAKA, Soichi WAKATSUKI, Koichi KATO 5* Nagoya City University, Nagoya, Aichi 467-8603, Japan, Tokyo Metropolitan Institute of Medical Science, Setagaya, Tokyo 156-8506, Japan as discussed by the authors, Photon Factory, KEK, Tsukuba,
Abstract: Eri SAKATA , Tadashi SATOH , Shunsuke YAMAMOTO, Yoshiki YAMAGUCHI , Maho YAGI-UTSUMI , Eiji KURIMOTO, Keiji TANAKA, Soichi WAKATSUKI, Koichi KATO 5* Nagoya City University, Nagoya, Aichi 467-8603, Japan, Tokyo Metropolitan Institute of Medical Science, Setagaya, Tokyo 156-8506, Japan, Photon Factory, KEK, Tsukuba, Ibaraki 305-0801, Japan, RIKEN Advanced Science Institute, Wako, Saitama, 351-0198, Japan, National Institutes of Natural Sciences, Okazaki, Aichi, 444-8787, Japan
01 Jan 2010
TL;DR: In this paper, the crystal structures of an intermediate of an UbcH5b ∼ ubiquitin (Ub) conjugate and Lys48-linked cyclic tetraubiquitin were reported.
Abstract: Ubiquitination is one of the most versatile protein modifications, which regulates a variety of cellular events. Here we report the crystal structures of an intermediate of UbcH5b ∼ ubiquitin (Ub) conjugate and Lys48-linked cyclic tetraubiquitin. These structures provide structural insights into the mechanisms underlying formation and elongation of the ubiquitin chains.