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Biomedical subjects

S Tanimoto

Publications and source records attributed to S Tanimoto.

At least 19 recordsLinked to original sources

Structural and functional properties of hen egg-white lysozyme deamidated by protein engineering.

The structural and functional properties of lysozymes genetically deamidated at positions 103 (N103D) and 106 (N106D) were studied by a protein engineering technique. The wild-type and mutant lysozymes were expressed in Saccharomyces cerevisiae and purified from the cultivation medium in two steps by cation-exchange chromatography on CM-Toyopearl. The lytic activity of deamidated lysozymes was almost the same as that of wild lysozyme, although the optimal pH of activity was slightly shifted to lower pH by the deamidation. The Gibbs free energy changes of unfolding (delta G) at 20 degrees C for N103D and N106D were almost the same as that of wild-type. On the other hand, the structural flexibility of lysozymes, estimated by protease digestion, was significantly increased by the deamidation. The surface functional properties of deamidated lysozymes were considerably enhanced, compared to those of wild-type lysozyme. These results suggest that structural flexibility is an important governing factor in surface functional properties of proteins, regardless of their structural stability.

Animals

Effects of static magnetic fields on diffusion in solutions.

Static magnetic fields affect the diffusion of biological particles in solutions through the Lorentz force and Maxwell stress. These effects were analyzed theoretically to estimate the threshold field strength for these effects. Our results show that the Lorentz force suppresses the diffusion of charged particles such as Na+, K+, Ca2+, Cl-, and plasma proteins. However, the threshold is so high, i.e., more than 10(4) T, that the Lorentz force does not affect the ion diffusion at typical field strengths (a few Tesla at most). Since the threshold of gradient fields for producing a change in ion diffusion through the Maxwell stress is more than 10(5) T2/m for paramagnetic molecules (FeCl3, O2) and plasma proteins, their diffusion would be unaffected by typical gradient fields (100 T2/m at most) and even by high gradient fields (less than 10(5) T2/m) used in magnetic separation techniques. In contrast, movement of deoxygenated erythrocytes and FeCl3 colloids (more than 10(3) molecules) is influenced by the usual gradient fields due to a volume effect.

Computer Simulation

Monoclonal antibodies to cowpox virus: polypeptide analysis of several major antigens.

Monoclonal antibodies directed against major antigens induced by cowpox virus (CPV) were produced. The specificities of these antibodies were established by immunoprecipitation, immunoblotting and several serological analyses, and from the cross-reactivities of these antibodies with cells infected with various other poxviruses, ectromelia virus (EV), vaccinia virus and Shope fibroma virus. The antibodies defined included ones reacting with each of the known major antigens of poxviruses, i.e. the common antigen of all poxviruses (probably NP antigen), the Orthopoxvirus-specific antigen (probably LS antigen), the haemagglutinin, the cell surface antigen, the common A-type inclusions in CPV and EV, and the antigen involved in neutralization.

Animals

The characteristics of luteinizing hormone and its beta-subunit secretion in dispersed rat anterior pituitary cells.

Secretion of luteinizing hormone (LH) and its beta-subunit (LH-beta) by luteinizing hormone-releasing hormone (LH-RH) were studied in perifused rat anterior pituitary cells. LH release was dose related between 10(-10) and 10(-6)M LH-RH. Every 5-minute pulse of 10(-8)M LH-RH induced almost similar response to the initial LH release; neither the self-priming effect nor desensitization were significant. Continuous administration of 10(-8)M LH-RH caused an initial LH release, followed by distinct desensitization, which could not be overcome by subsequent pulses of 10(-8)M LH-RH. The results were quite similar in LH-beta release. The perifusion study is shown to be useful in analyzing the characteristics of LH and LH-beta release, and the present results suggest that LH-RH receptor-mediated regulation may be also involved in LH-beta release.

Animals

Polypeptide analysis with monoclonal antibodies of A type inclusion bodies induced by cowpox virus.

The antigen in A type inclusion bodies (ATIB) induced by cowpox virus (CPV) was examined by immunofluorescent staining with monoclonal antibodies and polypeptide analysis. In the immunofluorescence (IF) test, these monoclonal antibodies reacted only with cytoplasmic inclusion bodies in cells infected with CPV. The fluorescence became detectable in the cells 6-9 hours after infection with CPV. No fluorescence was detectable in cells infected with CPV in the presence of cytosine-I-beta-D-arabinofuranosyl-HCl (Ara C) or in cells infected with other poxviruses, such as vaccinia virus (VV) or Shope fibroma virus (SFV). On Western blotting and immunoprecipitation followed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), only one component with a molecular weight of about 160,000 (160 K) was detected in CPV-infected cells. This 160 K polypeptide was first detectable 12 hours after infection of cells with CPV, and was not detectable in infected cells in the presence of Ara C. The 160 K polypeptide was not detected in cells infected with VV or SFV, or in virions purified from CPV-, VV- or SFV-infected cells.

Animals

Cross-reactivity among cowpox, ectromelia and vaccinia viruses with monoclonal antibodies recognizing distinct antigenic determinants in A-type inclusion bodies.

Several monoclonal antibodies recognizing distinct antigenic determinants in A-type inclusion bodies (ATIB) induced by cowpox virus (CPV) were obtained to examine the cross-reactivity among various strains of poxviridae, comprising CPV, ectromelia virus (EV), vaccinia virus (VV) and Shope fibroma virus (SFV). The monoclonal antibodies were classified into at least 3 groups on the basis of the results of an immunofluorescence test and immunoblotting; i. e., strain-specific, CPV and EV-specific and Orthopoxvirus (CPV, EV and VV)-specific antibodies. Differences were found between the antigenic determinants of ATIB of LB strains (LB red and LB white) and other strains (Amsterdam, 53, 58 and 60) of CPV and also between those of ATIB of CPV and EV. An interesting finding was that VV also produces the antigen analogous to that associated with ATIB in CPV- and EV-infected cells despite the absence of morphologically defined ATIB.

Antibodies, Monoclonal

[Study on the development of the radioimmunoassay of the corticotropin releasing factor (CRF)].

Synthetic ovine CRF was conjugated to bovine serum albumin (BSA) with glutaraldehyde and used as an immunogen to generate antibodies in rabbits. One of the antisera revealed 40% binding with 125I-CRF at 1 : 12,000 dilution in RIA. The antiserum did not show any significant crossreaction with other hormones. The Chloramine-T method was used to label CRF with 125I and purified by Sephadex G-50 fine (1 X 103 cm) chromatography with 0.03 M phosphate buffer (pH 7.2) containing 0.1% BSA, 20mM EDTA, 200 KIU/ml Trasyrol was used for the RIA buffer and the separation of free and antibody-bound CRF was performed by the Dextran-Charcoal method. The tentative minimum sensitivity of the assay was 20-100 pg/tube. The dilution curve of 125I-CRF in the hypothalamus of rat was parallel to that of the standard. Immunoreactive CRF content in the hypothalamus of male, female and post ovariectomized rats was measured by RIA. The content in those of male and post ovariectomized rats was significantly higher than in female rats. These data indicate that the pituitary-adrenal axis may have some effects on post-menopausal status.

Animals