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Biomedical subjects

Koichi Kato

Publications and source records attributed to Koichi Kato.

At least 19 recordsLinked to original sources
PubMed2007

Comparison of the methods for profiling glycoprotein glycans--HUPO Human Disease Glycomics/Proteome Initiative multi-institutional study.

Mass spectrometry (MS) of glycoproteins is an emerging field in proteomics, poised to meet the technical demand for elucidation of the structural complexity and functions of the oligosaccharide components of molecules. Considering the divergence of the mass spectrometric methods employed for oligosaccharide analysis in recent publications, it is necessary to establish technical standards and demonstrate capabilities. In the present study of the Human Proteome Organisation (HUPO) Human Disease Glycomics/Proteome Initiative (HGPI), the same samples of transferrin and immunoglobulin-G were analyzed for N-linked oligosaccharides and their relative abundances in 20 laboratories, and the chromatographic and mass spectrometric analysis results were evaluated. In general, matrix-assisted laser desorption/ionization (MALDI) time-of-flight MS of permethylated oligosaccharide mixtures carried out in six laboratories yielded good quantitation, and the results can be correlated to those of chromatography of reductive amination derivatives. For underivatized oligosaccharide alditols, graphitized carbon-liquid chromatography (LC)/electrospray ionization (ESI) MS detecting deprotonated molecules in the negative ion mode provided acceptable quantitation. The variance of the results among these three methods was small. Detailed analyses of tryptic glycopeptides employing either nano LC/ESI MS/MS or MALDI MS demonstrated excellent capability to determine site-specific or subclass-specific glycan profiles in these samples. Taking into account the variety of MS technologies and options for distinct protocols used in this study, the results of this multi-institutional study indicate that MS-based analysis appears as the efficient method for identification and quantitation of oligosaccharides in glycomic studies and endorse the power of MS for glycopeptide characterization with high sensitivity in proteomic programs.

Carbohydrate Conformation↗
PubMed2007

Direct interactions between NEDD8 and ubiquitin E2 conjugating enzymes upregulate cullin-based E3 ligase activity.

Although cullin-1 neddylation is crucial for the activation of SCF ubiquitin E3 ligases, the underlying mechanisms for NEDD8-mediated activation of SCF remain unclear. Here we demonstrate by NMR and mutational studies that NEDD8 binds the ubiquitin E2 (UBC4), but not NEDD8 E2 (UBC12). Our data imply that NEDD8 forms an active platform on the SCF complex for selective recruitment of ubiquitin-charged E2s in collaboration with RBX1, and thereby upregulates the E3 activity.

Carrier Proteins↗
PubMed2007

Antibody arrays for quantitative immunophenotyping.

Detection of multiple surface antigens expressed on living cell is an important step for cell processing and clinical diagnosis. Here we describe the preparation of antibody arrays that allow parallel detection of multiple surface antigens through affinity binding of living cells. An antibody array was fabricated by photo-assisted patterning of an alkanethiol monolayer formed on a gold-coated glass plate and subsequent immobilization of antibodies specific for cell surface antigens in an array format. We demonstrate here that rapid phenotyping can be performed on the array for both adhesion-dependent and non-dependent cells by direct cell binding assays. The density of bound cells on each antibody spot was in accordance with their contents in an original suspension. This result suggests the feasibility of the array-based method for quantitative assessment of multiple antigen expression. These findings will serve to extend the range of fundamental and clinical applications of antibody arrays.

Antigen-Antibody Complex↗
PubMed2006

Interaction of N-linked glycans, having multivalent GlcNAc termini, with GM3 ganglioside.

GM3 ganglioside interacts specifically with complex-type N-linked glycans having multivalent GlcNAc termini, as shown for (1) and (2) below. (1) Oligosaccharides (OS) isolated from ConA-non-binding N-linked glycans of ovalbumin, whose structures were identified as penta-antennary complex-type with bisecting GlcNAc, having five or six GlcNAc termini (OS B1, B2), or bi-antennary complex-type having two GlcNAc termini (OS I). OS I is a structure not previously described. (2) Multi-antennary complex-type N-linked OS isolated from fetuin, treated by sialidase followed by beta-galactosidase, having three or four GlcNAc termini exposed. These OS, conjugated to phosphatidylethanolamine (PE), showed clear interaction with (3)H-labeled liposomes containing GM3, when various doses of OS-PE conjugate were adhered by drying to multi-well polystyrene plates. Interaction was clearly observed only with liposomes containing GM3, but not LacCer, Gb4, or GalNAcalpha1-3Gb4 (Forssman antigen). GM3 interaction with PE conjugate of OS B1 or B2 was stronger than that with PE conjugate of OS I. GM3 interacted clearly with PE conjugate of N-linked OS from desialylated and degalactosylated fetuin, but not native fetuin. No binding was observed to cellobiose-PE conjugate, or to OS-PE conjugate lacking GlcNAc terminus. Thus, GM3, but not other GSL liposomes, interacts with various N-linked OS having multiple GlcNAc termini, in general. These findings suggest that the concept of carbohydrate-to-carbohydrate interaction can be extended to interaction of specific types of N-linked glycans with specific GSLs. Natural occurrence of such interaction to define cell biological phenomena is under investigation.

Animals↗
PubMed2006

Hydrogen bonding makes a difference in the rhodium-catalyzed enantioselective hydrogenation using monodentate phosphoramidites.

A new generation of monodentate phosphoramidite ligands bearing a primary amine moiety was found to display comparable or better efficiency than bisphosphines in the Rh-catalyzed asymmetric hydrogenation of challenging substrates, such as (Z)-methyl alpha-acetoxyacrylate or (E)-beta-aryl itaconate derivatives, affording the corresponding hydrogenation products with excellent enantioselectivities (up to >99% ee). The presence of intermolecular hydrogen bonding (HB) between two monodentate ligands in the catalyst was found to be critical for excellent catalyst performance. This finding provides a basis for design and development of further catalyst systems using this type of monodentate phosphoramidite ligands.

Catalysis↗
PubMed2006

Combinatorial protein display for the cell-based screening of biomaterials that direct neural stem cell differentiation.

Neural stem cell (NSC) has emerged as a potential source for cell replacement therapy following traumatic injuries and degenerative diseases of the central nervous system. However, clinical applications of NSC further require technological advances especially for controlling differentiation of NSC. This study aimed at developing biomaterials that serve to expand undifferentiated NSC or to induce cells with specific phenotypes. Our approach is to construct composite biomaterials that consist of extracellular matrix components and growth factors. In order to optimize matrix-growth factor combinations, we conducted the parallel and rapid screening of composite biomaterials through assays using cell-based arrays. The photo-assisted patterning of an alkanethiol self-assembled monolayer was employed to achieve site-addressable combinatorial immobilization of natural and synthetic matrices incorporated with growth factors including epidermal growth factor (EGF), ciliary neurotrophic factor (CNTF), nerve growth factor (NGF), and neurotrophin-3 (NT-3). NSC obtained from the rat embryonic striatum was cultured directly on the array to screen for cell adhesion, proliferation, and promotion of neuronal and glial specification. The results showed that the significant number of cells adhered to laminin-1, fibronectin, ProNectin, and poly(ethyleneimine). It was found that cells proliferated most extensively on a spot with immobilized EGF among the spots with different matrix-growth factor combinations. The results also showed that neuronal differentiation was promoted on the spots with immobilized NGF or NT-3, and astroglial differentiation with CNTF. Importantly, observed effects of growth factors were frequently altered depending on the type of co-immobilized matrices, suggesting synergic effects of adhesion and growth factor signals.

Adsorption↗
PubMed2006

Structural basis for recognition of the nonclassical MHC molecule HLA-G by the leukocyte Ig-like receptor B2 (LILRB2/LIR2/ILT4/CD85d).

HLA-G is a nonclassical MHC class I (MHCI) molecule that can suppress a wide range of immune responses in the maternal-fetal interface. The human inhibitory immune receptors leukocyte Ig-like receptor (LILR) B1 [also called LIR1, Ig-like transcript 2 (ILT2), or CD85j] and LILRB2 (LIR2/ILT4/CD85d) preferentially recognize HLA-G. HLA-G inherently exhibits various forms, including beta(2)-microglobulin (beta(2)m)-free and disulfide-linked dimer forms. Notably, LILRB1 cannot recognize the beta(2)m-free form of HLA-G or HLA-B27, but LILRB2 can recognize the beta(2)m-free form of HLA-B27. To date, the structural basis for HLA-G/LILR recognition remains to be examined. Here, we report the 2.5-A resolution crystal structure of the LILRB2/HLA-G complex. LILRB2 exhibits an overlapping but distinct MHCI recognition mode compared with LILRB1 and dominantly recognizes the hydrophobic site of the HLA-G alpha3 domain. NMR binding studies also confirmed these LILR recognition differences on both conformed (heavy chain/peptide/beta(2)m) and free forms of beta(2)m. Binding studies using beta(2)m-free MHCIs revealed differential beta(2)m-dependent LILR-binding specificities. These results suggest that subtle structural differences between LILRB family members cause the distinct binding specificities to various forms of HLA-G and other MHCIs, which may in turn regulate immune suppression.

Antigens, CD↗
PubMed2006

Probing dynamics and conformational change of the GroEL-GroES complex by 13C NMR spectroscopy.

Bacterial chaperonin GroEL with a molecular mass of 800 kDa was studied by (13)C NMR spectroscopy. Carbonyl carbons of GroEL were labeled with (13)C in an amino acid specific manner in order to reduce the number of signals to be observed in the spectrum. Combination of selective labeling and site-directed mutagenesis enabled us to establish the sequence specific assignment of the (13)C resonances from GroEL. ADP-binding induced a chemical shift change of Tyr478 in the equatorial domain and His401 in the intermediate domain, but little of Tyr203 in the apical domain. Upon complex formation with co-chaperonin GroES in the presence of ADP, Tyr478 exhibits two peaks that would originate from the cis and trans rings of the asymmetric GroEL-GroES complex. Comparison between the line width of the GroEL resonances and those from GroES in complex with GroEL revealed broadening disproportionate to the size of GroEL, implying the existence of conformational fluctuations which may be pertinent to the chaperone activity. Based on these results, we concluded that (13)C NMR observation in combination with selective labeling and site-directed mutagenesis can be utilized for probing the conformational change and dynamics of the extremely large molecules that are inaccessible with current NMR methods.

Adenosine Diphosphate↗
PubMed2006

Clinical and radiological features related to the growth potential of meningioma.

Clinical and radiological features that help predict the growth potential of meningioma would be beneficial. The purpose of this study is to clarify the characteristics related to proliferating potential using the MIB-1 staining index. We analyzed the relationship of MIB-1 staining indices to characteristics of 342 consecutive patients with meningioma surgically removed between 1995 and 2004 by logistic regression analysis. One hundred and forty-nine of the patients with meningioma were >or=60 in age; 89 male; 48 recurrent; 203 symptomatic; 157 at the skull base; 124 over 20 cm(3); 24 multiple; 136 with edema; 117 with calcification. The MIB-1 staining index in 56 of 296 grade I meningiomas in WHO classification was >or= 3.0; in 27 of 28 grade II; and in 17 of 18 grade III, respectively. Logistic regression analysis demonstrated that male (odds ratio [OR], 2.374, p=0.003), recurrence (OR, 7.574, p=0.0001), skull base (OR, 0.540, p=0.035), calcification (OR, 0.498, p=0.019) were independent risk factors for a high MIB-1 staining index (>or=3.0); age, symptomatic, volume, multiple, edema were not. Male, recurrence, non-skull base, absence of calcification are independent risk factors for a high proliferative potential. These should be taken into consideration when managing meningiomas.

Adult↗
PubMed2006

Evolutionally conserved intermediates between ubiquitin and NEDD8.

The investigation of common structural motifs provides additional information on why proteins conserve similar topologies yet may have non-conserved amino acid sequences. Proteins containing the ubiquitin superfold have similar topologies, although the sequence conservation is rather poor. Here, we present novel similarities and differences between the proteins ubiquitin and NEDD8. They have 57% identical sequence, almost identical backbone topology and similar functional strategy, although their physiological functions are mutually different. Using variable pressure NMR spectroscopy, we found that the two proteins have similar conformational fluctuation in the evolutionary conserved enzyme-binding region and contain a structurally similar locally disordered conformer (I) in equilibrium with the basic folded conformer (N). A notable difference between the two proteins is that the equilibrium population of I is far greater for NEDD8 (DeltaG(0)(NI)<5 kJ/mol) than for ubiquitin (DeltaG(0)(NI)=15.2(+/-1.0) kJ/mol), and that the tendency for overall unfolding (U) is also far higher for NEDD8 (DeltaG(0)(NU)=11.0(+/-1.5) kJ/mol) than for ubiquitin (DeltaG(0)(NU)=31.3(+/-4.7) kJ/mol). These results suggest that the marked differences in thermodynamic stabilities of the locally disordered conformer (I) and the overall unfolding species (U) are a key to determine the functional differences of the two structurally similar proteins in physiology.

Amino Acid Sequence↗
PubMed2006

Peptide recognition: encapsulation and alpha-helical folding of a nine-residue peptide within a hydrophobic dimeric capsule of a bowl-shaped host.

A dimeric capsule of coordination bowl 1 encapsulated a nine-residue peptide (Trp-Ala-Glu-Ala-Ala-Ala-Glu-Ala-Trp; 2) within the large hydrophobic cavity in water, and stabilized the alpha-helical conformation of bound 2. An NMR titration experiment revealed that monomeric bowl 1 recognized two Trp residues at the both terminals of 2 through 1/2 = 1:1 to 2:1 complexation. The 1:1 and 2:1 species exist in equilibrium even in the presence of excess 1. It was found that the formation of the 2:1 complex, in which two bowls of 1 wrapped the whole of 2, became dominant by the addition of NaNO3 due to the fact that the enhanced ion strength increased the hydrophobic interaction between Trp residues and the cavity of 1. The alpha-helical conformation of 2 within the dimeric capsule of 1 was elucidated from detailed NOESY analysis.

Dimerization↗
PubMed2006

Apolipoprotein A-I increases association of cytosolic cholesterol and caveolin-1 with microtubule cytoskeletons in rat astrocytes.

Apolipoprotein (apo) A-I induces rapid translocation of protein kinase Calpha and phospholipase Cgamma, and slow translocation of caveolin-1 and newly synthesized cholesterol to the cytosolic lipid-protein particle (CLPP) fraction in rat astrocytes. In order to understand the function of CLPP, we investigated the interaction with cytoskeletons of CLPP-related proteins such as caveolin-1 and protein kinase Calpha and of CLPP-related lipids in rat astrocytes. Under the conditions that microtubules were depolymerized, association of cytosolic caveolin-1 with protein kinase Calpha and alpha-tubulin was enhanced when the cells were treated with apoA-I for 5 min. This association was suppressed by a scaffolding domain-peptide of caveolin-1. Association with the microtubule-like filaments of cytosolic lipids, caveolin-1 and protein kinase Calpha was also increased by the apoA-I treatment and inhibited by the scaffolding domain peptide. Paclitaxel (taxol), a compound to stabilize microtubules, suppressed the apoA-I-mediated intracellular translocation and release from the cells of the de novo synthesized cholesterol and phospholipid. The findings suggested that the association of CLPP with microtubules is mediated by a scaffolding domain of caveolin-1, induced by apoA-I and involved in regulation of intracellular cholesterol trafficking for assembly of cellular lipids to apoA-I-high-density lipoprotein (HDL).

Animals↗
PubMed2006

Edible bird's nest extract inhibits influenza virus infection.

Edible bird's nest (EBN) is the nest of the swift that is made from its saliva. Although EBN has been widely used for enhancing immunocompetence, its antiviral efficacy has not been studied in detail. We found that EBN extract could strongly inhibit infection with influenza viruses in a host range-independent manner when it was hydrolyzed with Pancreatin F. Western blotting assay showed that the EBN extract bound to influenza virus. Furthermore, EBN extract could neutralize the infection of MDCK cells with influenza viruses and inhibit hemagglutination of influenza viruses to erythrocytes, but it could not inhibit the activity of influenza virus sialidase. Fluorometric HPLC indicated that the major molecular species of sialic acid in EBN is N-acetylneuraminic acid. The results suggest that EBN is a safe and valid natural source for the prevention of influenza viruses.

Animals↗
PubMed2006

Solution structure and dynamics of Ufm1, a ubiquitin-fold modifier 1.

The ubiquitin-fold modifier 1 (Ufm1) is one of various ubiquitin-like modifiers and conjugates to target proteins in cells through Uba5 (E1) and Ufc1 (E2). The Ufm1-system is conserved in metazoa and plants, suggesting its potential roles in various multicellular organisms. Herein, we analyzed the solution structure and dynamics of human Ufm1 (hsUfm1) by nuclear magnetic resonance spectroscopy. Although the global fold of hsUfm1 is similar to those of ubiquitin (Ub) and NEDD8, the cluster of acidic residues conserved in Ub and NEDD8 does not exist on the Ufm1 surface. 15N spin relaxation data revealed that the amino acid residues of hsUfm1 exhibiting conformational fluctuations form a cluster at the C-terminal segment and its spatial proximity, which correspond to the versatile ligand-binding sites of Ub and other ubiquitin-like proteins (Ubls). We suggest that Ub and other Ubl-modifiers share a common feature of potential conformational multiplicity, which might be associated with the broad ligand specificities of these proteins.

Amino Acid Sequence↗
PubMed2006

Layer-by-layer assembly of cationic lipid and plasmid DNA onto gold surface for stent-assisted gene transfer.

Intravascular stent-assisted gene transfer is an advanced approach for the therapy of vascular diseases such as atherosclerosis and stenosis. This approach requires a stent that allows local and efficient administration of therapeutic genes to the target cells at the vascular wall. To create such a stent, a method was developed for loading plasmid DNA onto the metal surface. The method involves the formation of self-assembled monolayer on the noble metal surface followed by electrostatic layer-by-layer (LBL) assembly of a cationic lipid/plasmid DNA complex and free plasmid DNA. In this in vitro feasibility study, the thin plainer film and the wire of gold were used as a substrate. The LBL assembly process was characterized by surface plasmon resonance spectroscopy and static contact angle measurement. Plasmid DNA loaded in the multilayer exhibited improved resistance against nuclease digestion. When cultured directly on the DNA-loaded surface, cells were transfected to express exogenous gene in the DNA loading-dependent manner. Plasmid DNA could also be transferred to endothelial cells from its apical side by placing the DNA-loaded gold wire onto the cell layer.

Cations↗
PubMed2006

Solution structures and behavior of trans-RuH(eta(1)-BH(4)) (binap)(1,2-diamine) complexes.

The solution structures of a number of trans-RuH(eta(1)-BH(4))[(S)-tolbinap](1,2-diamine) precatalysts [TolBINAP = 2,2'-bis(di-4-tolylphosphino)-1,1'-binaphthyl; 1,2-diamine==(S,S)- or (R,R)-1,2-diphenylethylenediamine (DPEN), ethylenediamine (EN), and (S)-1,1-di(4-anisyl)-2-isopropylethylenediamine (DAIPEN)] have been determined using 2D NMR ((1)H--(1)H DQF-COSY, (1)H--(13)C HMQC, (1)H--(31)P HSQC, and (1)H--(15)N HSQC), and a double-pulsed field-gradient spin-echo (DPFGSE) NOE technique. All the octahedral Ru complexes adopt a trans configuration with respect to the BH(4) and hydride ligands. Amine protons of trans-RuH(eta(1)-BH(4))[(S)-tolbinap](1,2-diamine) complexes undergo H/D exchange in (CD(3))(2)CDOD. This inherent high acidity, coupled with the lability and chemical properties of the BH(4) ligand, allows for precatalyst activation without the need for an added base, in contrast to trans-RuCl(2)[(S)-tolbinap](1,2-diamine) precatalysts, which require a strong base for generation of a catalytic species. The H/BH(4) complex in a 2-propanol solution is converted to catalytically active [trans-RuH{(S)-tolbinap}{(S,S)-dpen}(ROH)](+) [(RO)(ROH)(n)](-) (R = (CH(3))(2)CH), a loosely associated ion pair of the discrete (solvated) cationic fragment and anionic species.

Amines↗