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Nobuhisa Watanabe

Publications and source records attributed to Nobuhisa Watanabe.

32 records · Page 2Linked to original sources
PubMed2004

Crystallization and preliminary X-ray diffraction study of the catalytic subunit of archaeal H+ -transporting ATP synthase from Pyrococcus horikoshii OT3.

H+ -transporting ATP synthase (H+ -ATPase) is a multi-subunit complex which acts to produce ATP molecules. The catalytic subunit A of the archaeal-type H+ -ATPase from Pyrococcus horikoshii OT3 was cloned, expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method with MPD as a precipitant. X-ray intensity data were collected to 2.55 A resolution at beamline BL41XU of SPring-8. The crystals belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell parameters a = b = 128.0, c = 104.7 A, and contain one molecule per asymmetric unit.

Catalytic Domain↗
PubMed2004

Structure analysis of PH1161 protein, a transcriptional activator TenA homologue from the hyperthermophilic archaeon Pyrococcus horikoshii.

The crystal structure of the Bacillus subtilis TenA-homologue protein PH1161 from the hyperthermophilic archaebacterium Pyrococcus horikoshii was determined. TenA is known to belong to a new family of activators that stimulate the production of extracellular proteases in B. subtilis. A sequence-similarity search revealed that TenA-homologue proteins are widespread in bacteria and archaea, suggesting that this family of proteins plays an essential role in these organisms. In the present study, the first three-dimensional structure of a member of the TenA family of proteins was determined, unexpectedly revealing that the protein has a fold identical to that of haem oxygenase-1. Analysis has also shown that the protein has a unique ligand-binding pocket. Electron density of a bound ligand molecule was observed in this pocket. These results provide a valuable insight into the functional understanding of the TenA family of proteins.

Amino Acid Sequence↗
PubMed2004

Congener-specific intake fractions for PCDDs/DFs and Co-PCBs: modeling and validation.

Intake fractions (iFs) for emissions to air, water, and soil for 17 PCDDs/DFs and 12 Co-PCBs were calculated with a level III multimedia model and a food-chain exposure model in succession. The two integrated models were tested by comparing the predicted and measured concentrations in the environment and by comparing intakes through food. Measurement-based iFs were also calculated and compared with the model-based iFs. The air concentrations predicted by the fate model were close to the median of the observed concentrations, whereas the predicted soil and water concentrations were one-third to one-tenth the observed concentrations. This difference was large in case of PCDDs and Co-PCBs, which was explained by the past pollution such as commercial PCB products and PCDD impurities in chloronitrofen (CNP) and pentachlorophenol (PCP). For fish, the predicted and observed exposures agreed well each other. For meat and milk, the predicted exposures were about 10 times the observed exposures for PCDDs/DFs, whereas the predicted and observed values agreed well for Co-PCBs. When the model was modified to consider feeding of fish meal to livestock and geographic bias in feed-grass production, the predicted congener profile was comparable to the measured profile. The comparison also suggested that chickens should be modeled separately from other terrestrial livestock. The model-based iFs for air emission of OCDD and 2378-TCDD were 0.001% and 0.1%, respectively. The iFs of most Co-PCBs were higher than those of PCDDs/DFs. These iF differences suggest the importance of the fate factor in assessing emissions of the 29 congeners.

Animal Feed↗
PubMed2004

Combustible and incombustible speciation of Cl and S in various components of municipal solid waste.

Chlorine (Cl) and sulfur (S) in municipal solid waste (MSW) are important reactive elements during combustion. They generate the acidic pollutants HCl and SOx, and, furthermore, produce and suppress organic chlorinated compounds. Nevertheless, few practical reports about Cl and S content in MSW have been published. In combustion and recycling processes, both combustible Cl and S, and incombustible Cl and S species are equally important. This paper presents the results of a comprehensive study about combustible and incombustible Cl and S in MSW components, including kitchen garbage, paper, textiles, wood and leaves, plastics and small chips. By integrating this collected data with data about MSW composition, not only the overall content of Cl and S in MSW, but also the origins of both combustible and incombustible Cl and S were estimated. The average Cl content in bulk MSW was 3.7 g/kg of raw MSW, of which 2.7 and 1.0 g/kg were combustible and incombustible, respectively. The Cl contribution from plastics was 76% and 27% with respect to combustible and incombustible states. The average S content in bulk MSW was 0.81 g/kg of raw MSW, of which 0.46 g/kg was combustible and 0.35 g/kg was incombustible. Combustible S was mainly due to synthetic textiles, while incombustible S was primarily from paper.

Chlorine↗
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↗
PubMed2002

Ile-Lys-Val-Ala-Val (IKVAV)-containing laminin alpha1 chain peptides form amyloid-like fibrils.

The Ile-Lys-Val-Ala-Val (IKVAV) sequence derived from laminin-1 promotes cell adhesion, neurite outgrowth, and tumor growth and metastasis. Here, we examined amyloid formation of an IKVAV-containing peptide (LAM-L: AASIKVAVSADR, mouse laminin alpha1 chain 2097-2108). The LAM-L peptide was stained with Congo red and exhibited fibrils in electron microscopy with a characteristic cross-beta X-ray diffraction pattern. Further, infrared spectra of LAM-L suggested a beta-sheet structure. These results indicate that LAM-L forms amyloid-like fibrils. We also examined amyloid-like fibril formation of LAM-L analogs. The neurite outgrowth activity of the LAM-L analogs was closely related to their amyloid-like fibril formation.

Amyloid↗
PubMed2002

Semi-automatic protein crystallization system that allows in situ observation of X-ray diffraction from crystals in the drop.

Semi-automatic protein crystallization device was developed using 96 format Hydra dispenser. Eight extra syringes were added for pipetting protein solution with its own dispensing mechanism. This device allows setting up 96 drops in about three minutes. New crystallization plate has also been developed. The plate allows collecting an oscillation photograph of the crystal directly in the drop. The diffraction quality of the crystals in the drops can be inspected without causing any other physical perturbations to the crystals. With the modified dispenser and this plate, we can perform both hanging and sitting drop vapour diffusion method.

Archaeal Proteins↗
PubMed2002

The crystal structure of human MRP14 (S100A9), a Ca(2+)-dependent regulator protein in inflammatory process.

Human MRP14 (hMRP14) is a Ca(2+)-binding protein from the S100 family of proteins. This protein is co-expressed with human MRP8 (hMRP8), a homologue protein in myeloid cells, and plays an indispensable role in Ca(2+)-dependent functions during inflammation. This role includes the activation of Mac-1, the beta(2) integrin which is involved in neutrophil adhesion to endothelial cells. The crystal structure of the holo form of hMRP14 was analyzed at 2.1 A resolution. hMRP14 is distinguished from other S100 member proteins by its long C-terminal region, and its structure shows that the region is extensively flexible. In this crystal structure of hMRP14, Chaps molecules bind to the hinge region that connects two EF-hand motifs, which suggests that this region is a target-binding site of this protein. Based on a structural comparison of hMRP14 with hMRP8 and human S100A12 (hS100A12) that is another homologue protein, the character of MRP8/14 hetero-complex and the functional significance of the flexibility of the C-terminal region of hMRP14 are discussed.

Amino Acid Sequence↗
PubMed2002

The X-ray crystal structure of pyrrolidone-carboxylate peptidase from hyperthermophilic archaea Pyrococcus horikoshii.

The crystal structure of pyrrolidone-carboxylate peptidase (PCP) from hyperthermophilic archaea Pyrococcus horikoshii (PhoPCP) has been determined at 1.6-A resolution by X-ray crystallography. PCP belongs to the C15 family of cysteine protease, and specifically removes the amino terminal pyroglutamate residue from a wide range of N-terminal-blocking peptides. The crystal structure is very similar to that of other hyperthermophiles, Pyrococcus furiosus and Thermococcus litoralis, and even that from the mesophile, Bacillus amyloliquefaciens. The inter-subunit disulfide bonds, which have been proposed as one of the thermostabilizing factors of the PCP from such hyperthermophiles, was not present in PhoPCP. The result suggests that the thermostability of PhoPCP may be obtained by the accumulation of many weak factors.

Amino Acid Sequence↗
PubMed2002

Determination of fluorine, chlorine, bromine and iodine by barrier discharge radiofrequency helium plasma.

A determination method of fluorine (F), chlorine (Cl), bromine (Br) and iodine (I) by a barrier discharge radiofrequency helium plasma-atomic emission spectroscopy was developed. A borosilicate glass was wrapped by two copper film electrodes, one of which was earthed, and the other was supplied with a radiofrequency high voltage (98 kHz, 3.2 kV), resulting in a discharge inside of the tube. An optical emission from the discharge tube was introduced to a charge-coupled device (CCD)-spectrometer through an optical fiber, and was monitored in the wavelength range of 730-960 nm. The emission lines of F (733.2 nm, 739.9 nm), Cl (833.3 nm, 837.6 nm, 858.6 nm, 894.8 nm, 912.1 nm, etc.), Br (827.2 nm, 882.5 nm, 889.8 nm, 926.5 nm, etc.) and I (905.8 nm) were observed. The linearity of the calibration was determined for F and Cl over the range of 1-10 microg, and for Br of 0.1-1 microg. The relative emission intensity was in the order of Br > I > Cl >F.

Journal Article↗