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Biomedical subjects

K Takeo

Publications and source records attributed to K Takeo.

At least 73 records · Page 4Linked to original sources

Crystal structure of methyl 3-O-beta-D-glucopyranosyl-beta-D-glucopyranoside (methyl beta-D-laminarabioside) monohydrate.

Crystals of methyl 3-O-beta-D-glucopyranosyl-beta-D-glucopyranoside (methyl beta-D-laminarabioside) belong to the orthorhombic system, space group P2(1)2(1)2, with a = 14.548(2), b = 24.252(7), c = 4.938(1) A, and Z = 4. The crystal structure was solved by the direct method and refined by the full-matrix least-squares procedure to an R-value of 0.062 for 1099 observed reflections in the X-ray data. The molecular structure is similar to that of beta-D-laminarabiose. The torsional angles around the glycosidic bonds are influenced by both the existence of an intramolecular hydrogen bond at O-4'... O-5 and the exo-anomeric effect. In the nonreducing residue, the exocyclic O-5-C-5-C-6-O-6 and C-4-C-5-C-6-O-6 torsional angles are (+)gauche and trans, respectively, whereas the corresponding torsional angles in the reducing residue are (-)gauche and (+)gauche. One water molecule cocrystallizes with each disaccharide, and the crystal structure is stabilized mainly by intra- and inter-molecular hydrogen bonds involving water molecules.

Carbohydrate Conformation↗

Analysis of the interaction between an alpha (1----6)dextran-specific mouse hybridoma antibody and dextran B512 by affinity electrophoresis.

Carbohydrates are common environmental antigens. As dextran B512 is composed of a repeating structure of simple antigenic determinants, it is widely used to study the immunochemical properties of immunoglobulins. Two-dimensional affinity electrophoresis patterns of a mouse monoclonal antidextran antibody (35.8.2H; IgG1, BALB/c) were produced to obtain insights into the microheterogeneity of the monoclonal antibody. The monoclonal antibody was separated into about six spots which had an identical affinity to dextran B512, but differed in their isoelectric points (pI). In addition, the pH dependence of the binding affinity of this antidextran to dextran B512 was examined. By comparing affinities obtained by affinity electrophoresis between weakly basic (pH 9.5) and weakly acidic (pH 3.8) discontinuous buffer systems, the latter showed an affinity about 500 times lower than the former. The change in the affinity was investigated with a continuous pH gradient by an affinity titration curve and was seen to change markedly at about pH 6. This suggests that the histidine at residue 34 in the light-chain CDR1 is largely responsible for the dextran binding.

Amino Acid Sequence↗

Characterization of the interaction between human plasma fibronectin and collagen by means of affinity electrophoresis.

The interaction between human plasma fibronectin and different types and forms of collagen were analysed by affinity electrophoresis at different pH values. The fibronectin bound tightly to collagen type I, III and IV, but not to type V. The fibronectin interacted better with the denatured form of collagen type I (gelatin) than with the native form. At pH less than 5.5 the fibronectin exhibited much lower affinity to gelatin than at pH greater than 8.0. The interaction between the fibronectin and gelatin was further analysed by affinity electrophoresis in which apparent dissociation constants (Kd) of the fibronectin for gelatin were calculated, and effects of urea, 2-mercaptoethanol and temperature on the interaction were examined. The fibronectin markedly diminished its affinity to gelatin at 3 M urea to give Kd = 2.5 x 10(-6) M, which was 1000 times larger than the value without urea. The fibronectin dissociated into its monomers and the monomers diminished their affinity to gelatin in a stepwise fashion with increase in concentration of 2-mercaptoethanol. The fibronectin diminished the affinity to gelatin by elevating temperature, and van't Hoff plots of log Kd values against the reciprocal of absolute temperature (T) showed that log Kd was inversely proportional to 1/T in the range 15-50 degrees C, and the thermodynamic parameters of the standard enthalpy change, the standard free energy change and the entropy change at 37 degrees C for association of fibronectin and gelatin were all negative. At 60 degrees C the affinity of fibronectin to gelatin was not detectable.(ABSTRACT TRUNCATED AT 250 WORDS)

Collagen↗

Separation of monoclonal antibodies from antihapten antisera by two-dimensional affinity electrophoresis.

A high-resolution two-dimensional affinity electrophoresis (2D-AEP) method was developed, using capillary polyacrylamide gel (PAG) isoelectric focusing in the first and slab PAG affinity electrophoresis in the second direction. Using this method, anti-hapten antibodies were separated into a number of monoclonal antibody [immunoglobulin G (IgG)] families, each of which is composed of several IgG spots having an identical affinity to the hapten but different isoelectric points. 2D-AEP may offer a powerful tool for solving fundamental problems in immunochemistry such as antibody heterogeneity, its hapten binding specificity and antigen-dependent somatic mutation.

Antibodies, Monoclonal↗

Synthesis of nigero-oligosaccharides.

Nigerose [alpha-D-Glcp-(1----3)-D-Glcp], nigerotriose, nigerotetraose, and nigeropentaose have been synthesized by chain elongation starting at the reducing end, from the corresponding octa-, undeca-, tetradeca-, and heptadeca-beta-D-acetates, respectively, via thioglycoside-mediated 1,2-cis coupling, using 1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose as the glucosyl acceptor and methyl 2,3,4,6-tetra-O-benzyl-1-thio-beta-D-glucopyranoside, methyl 3-O-allyl-2,4,6-tri-O-benzyl-1-thio-beta-D-glucopyranoside, and methyl O-(2,3,4,6-tetra-O-benzyl-alpha-D-glucopyranosyl)-(1----3)-2,4,6-tri-O- benzyl-1-thio-beta-D-glucopyranoside as the donors.

Carbohydrate Sequence↗

Analysis of collagen type III by uninterrupted sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting: changes in collagen type III polymorphism in aging rats.

A new method of type III collagen analysis by uninterrupted sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) combined with immunoblotting was developed. The electrophoresis was carried out with gels containing 4 M urea. A negatively charged reducing agent, thioglycolic acid, was added to the running buffer of the cathodic reservoir between 15 and 20 min after Bromphenol Blue (BPB) migrated to the top of the separating gel, to reduce interchain disulfide binding of the collagen. The polymorphic type III collagens, i.e., an alpha-chain derived from a trimer [alpha 1(III)]3 with interchain disulfide bonds but without covalent cross-links, alpha 1(III), a beta-chain with covalent cross-links, beta(III), or an alpha-chain released from a trimer without reduction of the disulfide bonds, alpha*1(III), were identified by immunostaining and quantified by densitometry. Using this method, changes in collagen type III polymorphism with aging were examined in the aorta, brachial artery, and skin of rats. The total quantity of collagen type III decreased with aging in all tissues. beta(III) was the major component in the aorta and brachial artery, but alpha 1(III) was the major component in the skin. With increasing age from 3 to 60 weeks, the ratio of beta(III) to alpha 1(III), which is correlated with the extent of covalent cross-linking, showed a steep increase in the aorta but only a slight increase in the skin and it remained almost constant in the brachial artery.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

Analysis of human tear proteins by two-dimensional electrophoresis.

Human tear proteins in the conjunctival sac were separated on the basis of the differences in their isoelectric points and molecular weights using micro two-dimensional electrophoresis combined with immunoblotting. The two-dimensional electrophoretic patterns of tear proteins from patients with conjunctivitis were compared with those from normal individuals. We also measured integrated intensities of seven protein spots, lactoferrin (LF), albumin and five specific tear proteins (STP), to examine differences in the amounts of these proteins in tears from normal individuals of different sexes. In the tears from patients with conjunctivitis, secretory immunoglobulin A (IgA), LF and STP spots were stained more weakly, whereas the albumin spot was stained more strongly as compared with those from normal individuals. Furthermore, haptoglobin and IgG spots appeared in the tears from patients with conjunctivitis. These were more prominent in the tears from patients with severe conjunctivitis. There were significant differences in the amounts of LF and two kinds of STPs in the different sexes. The amounts of these proteins were larger in females.

Adult↗

Synthesis of the oligosaccharides alpha-D-Glcp-(1----4)-D-Xylp, alpha-D-Xylp-(1----4)-D-Glcp, alpha-D-Glcp-(1----4)-alpha-D- Glcp-(1----4)-D-Xylp, alpha-D-Glcp-(1----4)-alpha-D-Xylp-(1----4)- D-Glcp, and alpha-D-Xylp-(1----4)-alpha-D-Glcp-(1----4)-D-Glcp.

Syntheses are described of the disaccharides, alpha-D-Glcp-(1----4)-D-Xylp, alpha-D-Xylp-(1----4)-D-Glcp, and alpha-D-Xylp-(1----4)-D-Xylp, and the trisaccharides, alpha-D-Glcp-(1----4)-alpha-D-Glcp-(1----4)-D-Xylp, alpha-D-Glcp-(1----4)-alpha-D-Xylp-(1----4)-D-Glcp, and alpha-D-Xylp-(1----4)-alpha-D-Glcp-(1----4)-D-Glcp as analogues of glucoamylase and soybean beta-amylase substrates.

Carbohydrate Conformation↗

Thermally induced conformational transition of double-stranded xanthan in aqueous salt solutions.

The thermally induced conformational transition of double-stranded xanthans (degree of pyruvate substitution, DSp = 0.45) having Mw = 3.1, 5.7, and 20.3 x 10(5) has been studied in aqueous salt solutions by high-sensitivity differential scanning calorimetry (DSC). The double strandedness of these samples in the ordered conformation was ascertained by the value of mass per unit length, ML = 2090 +/- 270 g mol-1 nm-1, which was determined from the contour length obtained by electron microscopic observations and the molecular weight by light scattering measurements. The temperature at half completion of the transition T 1/2 for these samples increased linearly with the logarithm of the cation (Na+, K+) concentration. The plot of 1/T1/2 vs the natural logarithm of cation (Na+) concentration in mM for the sample with Mw = 5.7 x 10(5) (15-SX) yielded the equation 10(3)/T1/2 = 3.45-0.159 ln [Na+]. The specific enthalpy delta hcal for 15-SX, essentially independent of salt concentration above 20 mM, was 8.31 +/- 0.39 J/g (SD, n = 6). No systematic dependence of molecular weight on the transition temperature and the enthalpy was observed. Application of the Manning polyelectrolyte theory to the system using the DSC data suggested that the separation of the double strand of xanthan into two single chains was not completed at the temperature where the endothermic peak was finished. This suggestion is consistent with recent findings by light scattering measurements as a function of temperature. Our DSC study was extended to include four other samples from various sources. It was found that T1/2 and delta hcal depend on the pyruvate contents of the samples. For example, the t1/2 (t1/2/degrees C = T1/2/K - 237.15) values for samples with high pyruvate content (DSp = 0.9) and depyruvated (DSp = 0.14) in 20 mM aqueous NaCl were 48.8 and 85.3 degrees C, respectively. Two other samples showed relatively broad DSC curves having shoulders, which were resolved into two independent components. Thermodynamic parameters for each component were examined as a function of salt concentration, and the results obtained were interpreted in terms of the heterogeneity of the pyruvate content of the samples.

Calorimetry, Differential Scanning↗

Analysis of the interaction between human plasma fibronectin and gelatin by affinity electrophoresis.

The interaction between human plasma fibronectin and gelatin was analyzed by affinity electrophoresis, in which the fibronectin was subjected to electrophoresis in a 4% polyacrylamide gel in the presence and absence of gelatin, as an affinity ligand, and the fibronectin band was stained by an immunoblotting method. The apparent dissociation constants (Kd) of fibronectin for gelatin were calculated from affinity plots based on the original affinity equation at different pHs, urea concentrations, and temperatures. The fibronectin exhibited much lower affinity in the presence of urea. The Kds at 37 degrees C were 1.49 X 10(-7) M, 2.50 X 10(-6) M, and 3.58 X 10(-6) M with 2 M, 3 M, and 4 M urea, respectively. The van't Hoff plots of Kd values against absolute temperature (T) showed that the value of log Kd decreased in proportion to the increase in the value of 1/T within the range of 15-50 degrees C. The standard enthalpy, the standard free energy change at 37 degrees C, and the entropy change at 37 degrees C for association were calculated to be -124.7 kJ/mol, -33.23 kJ/mol, and -295.1 J/mol/deg, respectively. These results suggest that a hydrophilic interaction, such as hydrogen bond or van der Waals interaction, plays an important role in the binding of plasma fibronectin to gelatin.

Adult↗

Reevaluation of the teleomorph of the genus Histoplasma by ubiquinone systems.

Ubiquinone systems of the genus Histoplasma and Blastomyces dermatitidis were examined and the relationship between the two genera, Emmonsiella and Ajellomyces, was discussed. Ubiquinone systems have been adopted as a useful taxonomic criteria at the generic level for fungi. We investigated ubiquinone systems of sexual and asexual strains of Histoplasma species and Blastomyces dermatitidis. All the strains of the former group had dihydrogenated ubiquinone-10(Q-10(H2] as the major isoprenologue, while the latter had ubiquinone-10(Q-10). From the result, it was concluded that the teleomorph of Histoplasma species should be brought back to Emmonsiella.

Ascomycota↗

Karyology and hyphal characters as taxonomic criteria in ascomycetous black yeasts and related fungi.

Mycelial development of seventy-three strains of black yeasts and related fungi were studied, and numbers of nuclei per hyphal cell were counted. Two main patterns were apparent in expanding hyphae, viz. (1) uninucleate expanding hyphal cells, septum formation strictly following mitosis, and (2) multinucleate, branched, aseptate hyphal tips, septa being formed in a later stage, leading to oligo- or uninucleate mature cells. Characteristic genera in the two groups are Exophiala and Aureobasidium, respectively. In Zasmidium and in some Ramichloridium species all mycelial cells are oligonucleate. The character is indicative for relationships at the family level in black yeasts.

Ascomycota↗

Karyology and the possible function of the dual conidia of Dissoconium (Hyphomycetes).

The morphological development and nuclear behaviour of two species of Dissoconium are described. The fungi are monokaryotic in all stages of their life cycle. Liberated pairs of conidia may form anastomoses, probably serving survival under hostile environmental conditions. The function of the one-celled conidia in D. dekkeri remains unexplained.

Cell Cycle↗

Usefulness of glycosylated recombinant human lymphotoxin for growth inhibition of human and murine solid tumors and experimental metastasis in mice.

We have examined the antitumor and antimetastatic effects of native-type, glycosylated recombinant lymphotoxin (LT) on human and murine tumors transplanted in mice. The results reported here are as follows: (a) The in vivo antitumor spectrum of LT is not coincident with the in vitro study, and it has a wide antitumor spectrum and substantially inhibits the growth of human solid tumors, (b) When both syngeneic and nude mice are transplanted with Meth A tumor, the significant growth-inhibitory effect of LT is obtained in syngeneic mice, but the effect is quite small in nude mice regardless of the routes; LT attains the same degree of effectiveness as that in syngeneic mice, but at an 8 to 16 times higher dose. Furthermore, the pretreatment with anti-asialo-GM1 antibody inhibits the antitumor effects of LT in syngeneic mice, (c) In the pulmonary metastasis model induced by i.v. injection of Meth A cells, a high preventive effect of LT is obtained by systemic administration in syngeneic mice, but not in nude mice. In addition, the pretreatment with anti-asialo-GM1 antibody completely prevents the antimetastatic effect of LT, but also blocks that effect of control mice without LT treatment. In conclusion, LT appears to be a potent cytokine against tumor growth and metastasis in vivo. The differences between nude and syngeneic mice suggest the involvement of host immunity in the expression of LT function.

Animals↗

Synthesis of O-alpha-D-glucopyranosyl-(1----4)-O-alpha -D-xylopyranosyl-(1----4)-O-alpha -D-xylopyranosyl-(1----4)-D-glucopyranose as a substrate analogue of alpha amylase.

The tetrasaccharide a-D-Glcp-(1----4)-a-D-Xylp-(1----4)-a-D-Xylp-(1----4)-D- Glcp (1) has been synthesized, as a substrate analogue of alpha amylase, by silver perchlorate-catalyzed glycosylation of benzyl 2,3,6-tri-O-benzyl-4-O-(2,3-di-O-benzyl-a-D-xylopyranosyl)-beta-D- glucopyranoside (30) with 2,3-di-O-benzyl-4-O-(2,3,4,6-tetra-O-benzyl-a-D- glucopyranosyl)-a-D-xylopyranosyl chloride or by methyl triflate-promoted condensation of 30 with methyl 2,3-di-O-benzyl-4-O-(2,3,4,6-tetra-O-benzyl-a-D-glucopyranosyl)-1-thio- beta-D-xylopyranoside, followed by removal of protecting groups of the resulting tetrasaccharide derivative 40.

Carbohydrate Sequence↗

Synergistic antitumor effect of glycosylated recombinant human lymphotoxin with human interferon-gamma on lymphotoxin-sensitive human tumor.

We examined the antitumor effect of glycosylated recombinant lymphotoxin (LT) in combination with human interferon-gamma (IFN-gamma) on human tumors transplanted into nude mice and compared it with that of tumor necrosis factor (TNF). The results were as follows: (i) The systemic administration of glycosylated LT combined with IFN-gamma produced a significant antitumor activity against HT-1080 fibrosarcoma, G-361 malignant melanoma, KB nasopharyngeal carcinoma, and ZR-75-1 breast carcinoma, all of which are relatively resistant to a single treatment with LT or IFN-gamma. The synergistic effect was also seen in LT-sensitive HeLa S3 tumors. The effect was observed after either i.v. or s.c. injection. (ii) In contrast, no synergistic or additive effect on HeLa S3 tumors was observed in the case of TNF combined with IFN-gamma. (iii) The serum half-life of glycosylated LT in tumor-bearing mice was about 22-fold longer than that of TNF. In conclusion, glycosylated LT, especially in combination with IFN-gamma, appears to be a potent cytokine against tumor growth in vivo compared with TNF. Its long serum half-life can result in a strong antitumor effect in combination with IFN-gamma in vivo.

Amino Acid Sequence↗