Biomedical subjects
Publications and source records attributed to N Kamiya.
SPring-8 is constructing most of its beamlines using a combination of standardized components. The structures of the beamlines are also standardized according to the light-source characteristics. Specifications of components as well as the unified method of assembly and alignment are described.
A bleomycin-binding protein (BLMA) produced by bleomycin-producing Streptomyces verticullus was crystallized in a form suitable for X-ray diffraction analysis using the vapor-diffusion method. Crystals were grown at pH 5.7, in 0.2 M NH4 actate and 0.1 M Na acetate, using 30% PEG 4000 as a precipitant. They belong to the orthorhombic system, with space group P21212, cell dimensions a = 54. 90, b = 67.94, c = 35.60 A, and one BLMA molecule in the asymmetric unit. The crystals diffract X-rays well and the diffraction intensity data was collected up to 1.5 A resolution with a merging R value of 0.054 at beamline 6B of the Photon Factory. The diffraction data set is 94% complete.
An operative case of localized mesothelioma of the pleura developed in a 17-year-old female was reported. She was admitted to our hospital complaining of right chest pain. A chest X-ray film showed a huge mass in the right lower field of the lung. After the embolization of the right inferior phrenic artery, which was the main feeder to the tumor, it was then successfully resected combined with the right diaphragm and the right lower lobe of the lung. Histologically, the tumor was diagnosed as benign localized mesothelioma (solitary fibrous tumor of the pleura). Further histological and immunohistochemical study revealed that it had developed from the connective tissue under the parietal mesothelium of the diaphragm. As chest roentgenograms had been undertaken during past two years, the tumor volume doubling time was calculated at 153 days. Despite the short tumor volume doubling time as like primary lung cancer, she is alive without recurrence 5 years after the operation.
The iron-containing nitrile hydratase (NHase) is a photoreactive enzyme that is inactivated in the dark because of persistent association with NO and activated by photo-dissociation of NO. The crystal structure at 1.7 A resolution and mass spectrometry revealed the structure of the non-heme iron catalytic center in the nitrosylated state. Two Cys residues coordinated to the iron were post-translationally modified to Cys-sulfenic and -sulfinic acids. Together with another oxygen atom of the Ser ligand, these modifications induced a claw setting of oxygen atoms capturing an NO molecule. This unprecedented structure is likely to enable the photo-regulation of NHase and will provide an excellent model for designing photo-controllable chelate complexes and, ultimately, proteins.
For evaluation of anti-hepatitis B virus (HBV) drugs, we have employed the HBV transgenic mouse in which virion-like particles can be assayed in the serum. Bispivaloyloxymethyl-9-(2-phosphonylmethoxyethyl)-adenine [bis (POM) PMEA] 100 mg/kg/day, 2',3'-dideoxy-3'-thiacytidine [(+-)-BCH189] 200 mg/kg/day and a placebo were orally administered to mice twice a day for 14 days. Anti-viral effects were monitored by checking the levels of serum HBV DNA by the semiquantitative polymerase chain reaction, HBsAg and HBeAg by enzyme immunoassay, and replicative intermediates in the liver by Southern blotting. As expected, decrease from the 10(0.5) to 10(3) copies of HBV DNA per microl of sera detected before the treatment to the undetectable level was evident for all five animals treated with bis(POM) PMEA 100 mg/kg/day. However (+-)-BCH189 200 mg/kg/day, which is known to act as the inhibitor of reverse transcriptase for HBV or HIV in vivo and in vitro, did not suppress HBV DNA levels in the transgenic mouse. Thus, we were able to detect the effects of anti-HBV drugs semi-quantitatively, and confirm differences in drug efficacy.
A diffraction-intensity data-collection system with synchrotron radiation X-rays utilizing the screenless Weissenberg technique and incorporating a large-format imaging plate is one of the most suitable apparatus for time-resolved protein crystallography with larger angle oscillations than hitherto described. The time resolution and data quality of the system have been tested using a tetragonal lysozyme crystal as a test sample in a flow-cell experiment at the bending-magnet beamline 18B at the Photon Factory, and a time resolution of 15 min is confirmed.
Immunohistochemical studies were performed in order to characterize glomus tumors. In all cases the glomus tumor cells stained positively for smooth muscle actin. In half of six cases, the glomus tumor cells weakly expressed desmin. They were not reactive with Ulex europaeus agglutinin-I and anti-Factor VIII-related antigen, whereas the only endothelial cells were reactive with them. The tumor cells as well as vascular endothelial cells amongst the tumor cells expressed CD34 in five of the cases.
A two-dimensionally tunable focusing monochromator has been developed for protein crystallography at high-energy undulator beamlines of third-generation synchrotron radiation facilities. This monochromator consists of a silicon wafer fabricated with an oblique-cut angle between the Bragg net plane and the crystal surface, and adhered onto a table-like copper block. The radii of curvatures are altered independently in two directions by expanding the spaces between the table legs. The versatilities of the meridional and sagittal curvatures were confirmed by X-ray experiments and three-dimensional shape measurements, respectively. The two-dimensional focusing ability was demonstrated using high-energy X-rays of 37.7 keV emitted from a bending-magnet source at the Photon Factory. A quasi-isotropic profile of converged X-rays was achieved near the focal position. The apparent gain of photon flux was 21. As a result of these excellent monochromator characteristics, a diffraction pattern of a hen egg-white lysozyme crystal was successfully obtained using high-energy X-rays.
We herein present a patient with Morgagni hernia which was diagnosed by magnetic resonance imaging (MRI). The patient had a progressively enlarging mass in the right cardiophrenic angle on chest roentgenogram. On computed tomography (CT) scans, the mass was revealed to have fat density and therefore was suspected to be either a lipoma or liposarcoma. MRI clearly demonstrated that the mass shadow was composed of omental fat herniating into the right thorax through the diaphragmatic hiatus. MRI is thus considered to be a useful noninvasive modality for the evaluation of lower anterior mediastinal masses demonstrating fat density on CT.
An active substance with antitumor activity (ARS2) was purified from the culture media of Chlorella vulgaris and found to be a glycoprotein with a molecular weight of 63,100 amu, as determined by matrix-assisted laser desorption/ ionization (MALDI) mass spectrometry. ARS2 contains 66.9% carbohydrate, mainly D-galactose, and 35.2% protein. The carbohydrate moiety has a beta-1,6-D-galactopyranose backbone, as determined by methylation analysis and 13C-NMR. Apparently, the protein moiety, whose 15 amino acid sequence at the NH2-terminus, we determined as DVGEAFPTVVDALVA, is necessary for the antitumor activity, as assessed by hydrazinolysis, periodate oxidation, and proteolysis.
A 70-year-old woman with no history of smoking was admitted to the hospital because of a persistent cough and pneumonia in the right lower lobe. Examination with a fiberoptic bronchoscope revealed an intratracheal polypoid tumor and a polypoid tumor almost completely obstructing the right bronchus B(8). Biopsies were done, and histological diagnosis of both tumors was squamous cell carcinoma. A sleeve resection of the trachea with end-to-end anastomosis and right lower lobectomy were done. Examination of specimens resected from both tumors revealed moderately differentiated squamous cell carcinoma. Synchronous double-primary cancer (both squamous cell carcinoma) of the lung and trachea in a non-smoker is very rare.
The diagnostic process for small solitary pulmonary nodules was retrospectively analyzed. Three hundred and forty-three patients, each with a solitary pulmonary lesion of less than 3 cm on chest radiography, were evaluated. The patients were seen at our hospital between September 1986 and December 1993. They consisted of 138 with malignant diseases (114 with lung cancer and 24 with metastatic tumors), and 205 with benign diseases. There were 146 patients (71%) with benign lesions of less than 2 cm. The diagnosis in 118 patients (86%) with malignant disease was made by lung biopsy. Of these 118 patients, 19 underwent biopsy of the lung through a thoracoscope or open lung biopsy via a thoracotomy. Benign diseases were diagnosed in 167 patients (81%) by various laboratory tests and imaging techniques, including chest radiography. Thinsection CT was done in 132 patients. In 72 of these 132 patients (16 with lung cancer, 23 with metastatic tumors, and 33 with benign tumors), a judgement regarding whether the lesion was malignant or not could not be made. Therefore, open lung biopsy was done in 5 patients. Twenty-four patients were only observed, 43 underwent biopsy, and 35 underwent surgery. A decision tree needs to be established to select therapeutic options for management of patients whose diagnoses remain indeterminate even after thin-section CT. Formulation of a decision tree based not only on clinical findings but also on the results of diagnostic imaging is necessary to improve clinical decision making regarding small solitary pulmonary nodules.
We report an unusual case of thymic Hodgkin's disease with cystic variants. The patient complained of nodules in his neck. Chest radiographs showed a mass with cystic lesions in the anterior mediastinum. Although Hodgkin's disease was suspected from pathologic examination of biopsy specimens of the neck nodules, the mediastinal and neck masses could not be precisely diagnosed until a median sternotomy was done. The surgically excised mediastinal masses revealed thymic Hodgkin's disease with cystic lesions. The patient has done well after chemotherapy and radiotherapy. Thymic Hodgkin's disease with cystic variants is very rare and only seventeen cases of this disease have been reported.
Marek's disease (MD) virus (MDV) serotype 1 (MDV1) pp38, an antigen associated with MD-transformed cells, has been identified as an MDV1 serotype-specific antigen by analyses using specific monoclonal antibodies (MAbs). In the present study, we determined the region in the MDV serotype 2 (MDV2) genome that has homology with the MDV1 pp38 gene. As a result of sequence analysis, it was determined that an open reading frame (ORF) existed in this region. Furthermore, Northern blot analysis and reverse transcriptase polymerase chain reaction confirmed the existence of an RNA transcript related to the ORF in MDV2-infected cells. Computer analyses revealed that MDV1 pp38 could be divided into two parts; one part was highly homologous and the other part was not homologous with the peptides deduced from the ORF of MDV2, indicating that the epitope recognized by a MDV1 pp38-specific MAb is located in the latter part.
A novel ordered assemblage of bacteriorhodopsin, a transmembrane protein functioning as a light-driven proton pump, is found in its three-dimensional crystal. Atomic force microscope images of the crystal surface reveal that spherical protein clusters with a diameter of approximately 50 nm are hexagonally close-packed. Electron micrographs of mechanically disintegrated crystals show that the inside of the protein cluster is filled with the mother liquor. The crystal is made up of hollow protein clusters. When disintegrated crystals are illuminated in the presence of a lipophilic anion, a significant alkalization of the external medium occurs. This result indicates that the protein cluster contains native lipids and that the cytoplasmic side of the protein faces the external medium. X-ray diffraction patterns and the observed diameter of the spherical shell suggest that approximately 200 bacteriorhodopsin trimers are aligned on a polyhedral surface lattice. Another remarkable feature of the spherical assemblies of bacteriorhodopsin is that they fuse with each other at low ionic strength and occasionally form a tubular or doughnut-like structure. The concept of membrane protein polymorphism is introduced on the basis of these observations, and it is used to describe the dynamic structure of some other biological membranes.
Recombinant hemagglutinin-neuraminidase (rHN) and fusion (rF) glycoproteins of virulent and avirulent strains of Newcastle disease virus (NDV) expressed by using baculovirus expression system were used to investigate their protective immunization effects in chickens. The efficacy of immunization with these recombinant proteins was evaluated by challenge infection. The chickens immunized with either rHN or rF protein of a virulent strain or rHN protein of an avirulent strain were completely protected from the lethal infection of virulent NDV. On the other hand, the rF protein of an avirulent strain, in which precursor F protein was not cleaved, showed lower protective effects. Significant levels of specific antibodies against respective proteins were detected in sera from survivors, whereas relatively lower levels of antibodies were found in chickens which were killed by challenge infection. These data indicate that either HN or F protein alone could induce protective immune responses and the cleavage of F protein might be important for its immunological potential.
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