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Biomedical subjects

T Tsuchiya

Publications and source records attributed to T Tsuchiya.

At least 685 records · Page 38Linked to original sources
PubMed1984

Characteristics of n-octyl beta-D-thioglucopyranoside, a new non-ionic detergent useful for membrane biochemistry.

n-Octyl beta-D-thioglucopyranoside, a new non-ionic detergent, was synthesized. Properties, and applicability to membrane proteins, of this detergent were investigated. The detergent was easily removed by dialysis. The solubilizing power of this detergent for Escherichia coli membrane proteins was similar to that of n-octyl beta-D-glucopyranoside, which has been widely used in membrane biochemistry. No inactivation of proteins was observed after the solubilization. n-Octyl beta-D-thioglucopyranoside was superior to n-octyl beta-D-glucopyranoside in that it was much more stable and could be synthesized at much lower cost.

Cell Membrane↗
PubMed1984

Dissipation of membrane potential of Escherichia coli cells induced by macromolecular polylysine.

Macromolecular polylysine caused a rapid and drastic decrease in the membrane potential of Escherichia coli cells, monitored with the uptake of the lipophilic cation, tetraphenylphosphonium cation (TPP+). Respiration of cells was not affected to a significant extent by polylysine. Polylysine treatment induced the efflux of K+ from cells, suggesting that an increase in permeability of cytoplasmic membrane may be responsible for the decrease in the membrane potential. The action of polylysine is discussed in comparison with that of colicin E1 which is known to dissipate the membrane potential of E. coli cells.

Cell Membrane Permeability↗
PubMed1984

Nucleotide sequence of the promoter region of the melibiose operon of Escherichia coli.

The nucleotide sequence of the promoter region of the melibiose operon of E. coli was determined. Consensus sequences for the -35 region, the Pribnow box and the binding site for cyclic AMP receptor protein were found in this region. The possible secondary structure of this DNA region was very similar to that of the promoter region of the lactose operon. A possible initiation ATG preceded by a Shine-Dalgarno sequence with proper spacing was present just downstream of the promoter region. The possible sequence of 52 amino acid residues in the NH2 terminus of the alpha-galactosidase were determined.

Amino Acid Sequence↗
PubMed1984

Nucleotide sequence of the melB gene and characteristics of deduced amino acid sequence of the melibiose carrier in Escherichia coli.

The nucleotide sequence of the melB gene coding for the melibiose carrier in Escherichia coli has been determined. The melibiose carrier is predicted to consist of 469 amino acid residues, resulting in a protein with a molecular weight of 52,029. The predicted carrier protein is highly hydrophobic (70% nonpolar amino acid residues). The hydropathic profile suggests that there are 10 long hydrophobic segments in the primary structure of the carrier protein. Most of them seem to traverse the membrane. Although the hydropathic profile of the melibiose carrier is similar to that of the lactose carrier as a whole, homology in the primary structure between the two carriers is very low. Furthermore, no homology in the nucleotide sequence is found in the structural genes for the two carriers. However, the nucleotide sequences of the intergenic regions are very similar between the melibiose operon and the lactose operon. There is a typical intercistronic regulatory sequence in the 3'-flanking region of the melB as well as in that of the lacY, which suggests the presence of another gene downstream of the melB.

Amino Acid Sequence↗
PubMed1984

Cotransport of proline and Li+ in Escherichia coli.

Uptake of Li+ induced by the addition of proline to a cell suspension of Escherichia coli was detected using an Li+-selective electrode. This Li+ uptake was inhibited by L-azetidine 2-carboxylic acid, a competitive inhibitor of the proline transport system. Thus, direct evidence for Li+-proline cotransport via the proline transport system was obtained. Kinetic parameters of the Li+ uptake were determined.

Azetidinecarboxylic Acid↗
PubMed1984

Sensitivity to bleomycin of JTC-11 cells modified by serum concentration and underlaid 3T3 cells.

The sensitivity to bleomycin of JTC-11 cells derived from Ehrlich ascites tumor was modified by both serum concentration and superinoculation on a sheet of Swiss 3T3 cells. The dose-survival curve of the JTC-11 cells was biphasic or triphasic, with a sensitive phase at lower doses (less than 10 micrograms/ml) and a resistant one at higher doses (more than 40 micrograms/ml). Sensitivity of JTC-11 cells to bleomycin decreased with increases in the serum concentration. When JTC-11 cells were superinoculated on the sheet of Swiss 3T3 cells, the sensitivity of JTC-11 cells to bleomycin did not change with increases in the serum concentrations.

Animals↗
PubMed1984

Comparison of F1's of oxidative phosphorylation from Escherichia coli and Salmonella typhimurium and demonstration of interchangeability of their subunits.

The peripheral membrane portion (SF1) of proton-translocating ATPase of Salmonella typhimurium and its alpha, beta, and gamma subunits were purified and compared with the same portion (EF1) from Escherichia coli. The alpha, beta, and gamma subunits of these F1's were found to be mutually interchangeable, and all possible combinations of the three subunits from EF1 and SF1 showed ATPase activity. Both F1's could bind functionally to the integral membrane part (F0) of either bacterium, suggesting that F0 and F1 are interchangeable in these two bacteria and thus that the two F1's are closely similar at the level of subunit structure. However, SF1 differed from EF1 in some enzymological properties such as its specific activity and susceptibilities to sodium dodecyl sulfate and methanol. The specific ATPase activity of EF1 was more than twice that of SF1, and hybrid enzymes containing the beta subunit of EF1 had higher activity than other hybrids. Amino acid analysis suggested that the primary structures of the alpha subunits of the two F1's are less homologous than those of the beta subunits. Thus, the primary structure of the alpha subunit may be more species specific than that of the beta subunit.

Amino Acids↗
PubMed1984

Physical and genetic characterization of the melibiose operon and identification of the gene products in Escherichia coli.

We have identified hybrid plasmids carrying the melibiose operon of Escherichia coli in a colony bank of Clarke and Carbon (Tsuchiya, T., Ottina, K., Moriyama, Y., Newman, M., and Wilson, T. H. (1982) J. Biol. Chem. 257, 5125-5128). Using one of the plasmids as a starting material, the DNA fragments containing the melibiose operon were recloned in a vector pBR322. Restriction maps were prepared, and several DNA segments were subcloned into pBR322. Genetic complementation tests and recombination analyses using those plasmids and melA- and melB- mutants as well as biochemical analyses of mel mutants transformed with those plasmids enabled us to determine the physical location of promoter, melA, and melB on the DNA segment. The size of the melAB region was about 3,000 base pairs. Gene products were identified using maxicells harboring plasmids carrying the melibiose operon. The apparent molecular weight of the alpha-galactosidase (coded by melA) was about 50,000 and that of the melibiose carrier (coded by melB) was about 31,000, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The melibiose carrier was also identified as a 30,000-dalton protein in reconstituted proteoliposomes which possessed melibiose transport activity.

Disaccharides↗
PubMed1984

Muscle stiffness changes during isometric contraction in frog skeletal muscle as studied by the use of ultrasonic waves.

In order to measure muscle stiffness changes with a high time resolution and with minimal disturbance to the contractile mechanism per se, we constructed an apparatus with which the propagation velocity of ultrasonic waves (MHz region) in the longitudinal or transverse direction was measured to serve as a measure of muscle stiffness. The longitudinal muscle stiffness started to increase on stimulation before the onset of isometric force, and reached a maximum before the peak twitch force. Analysis of experimental data indicated that, during an isometric tetanus, the increment of muscle longitudinal stiffness was about 6 X 10(7)N/m2, a value similar to those obtained by Truong (1974) and Ford et al. (1981) with sinusoidal vibrations (3 kHz) and length steps respectively. This suggests that the increment of muscle longitudinal stiffness during the activation of the contractile system results from the recruitment of an almost non-dispersive elastic component. In the case of transverse muscle stiffness, on the other hand, it started to decrease on stimulation before the onset of isometric force, and reached a minimum before the peak twitch force. Possible causes of this unexpected result is discussed in relation to the molecular mechanism of muscle contraction.

Animals↗
PubMed1984

Depletion of the reduced glutathione level in the liver and production of the mutagens in the intestine in the mice inducing hepatoma by feeding on a high level dose of sorbic acid.

Mutagenicity was detected using Salmonella typhimurium TA98 in the acidic components of the intestinal contents of mice, in which a high incidence of hepatoma had been reported due to feeding on a diet containing 15% sorbic acid (Ishizawa et al., 1980). Furthermore, the glutathione level in the liver of the 15% sorbic acid group was decreased to 40% of the amount found in controls after a 3-month feeding period, and this low level was maintained for long periods (12 months) until the end of the experiments. There was also a close correlation between the extent of depletion of the hepato-glutathione level and the concentration of sorbic acid added to the diet. Consequently, the hepatoma which developed in mice fed a 15% sorbic acid diet was considered to be induced both by the depletion of the hepato-glutathione level over the long periods, and by the gradual production of various mutagens in the intestine which were absorbed and transferred to liver to be metabolically activated.

Animals↗
PubMed1984

Polydispersity effects on head-to-tail polymerization of F-actin. A theoretical treatment.

A theoretical description of the release of labeled subunits of F-actin that undergoes head-to-tail polymerization utilizing random-walk theory is extended to the case of a finite-length polymer. An exact expression is given for the number of released subunits in a suitable form for numerical calculation. By comparing the results for a finite-length polymer with that for an infinite-length case, the limitation of the infinite-length approximation is readily known. The results for finite-length polymers are applied to systems with polydispersity. It is shown that the number of labeled subunits released from F-actin depends strongly on the distribution functions of the polymer lengths and the values of the transition probabilities of the random walkers.

Actins↗
PubMed1984

The characterization of invertebrate troponin C.

TNCs from lobster, mussel, and squid migrated with rabbit TNC at an apparent mol. wt of 18,000. Electrophoretic mobilities in the presence or absence of Ca2+ were compared: the electrophoretic mobility of rabbit TNC was greater in the presence of Ca2+ than it its absence, but all invertebrate TNCs tested migrated identically, whether Ca2+ was present or not. The Ca2+-binding capacity of invertebrate TNCs was only one Ca2+ ion per molecule. The alpha-helix contents in the presence or absence of Ca2+ were compared: rabbit TNC changed by a value of 16% and invertebrate TNCs by 4%. Antibodies to loligo TNC did not cross-react with rabbit TNC, but did interact with their molluscan TNCs.

Animals↗
PubMed1984

Solubilization and reconstitution of membrane proteins of Escherichia coli using alkanoyl-N-methylglucamides.

Alkanoyl-N-methylglucamides, nonionic detergents, were utilized to solubilize membrane proteins of Escherichia coli and were used to reconstitute them into liposomes. First, critical micelle concentrations (CMC) of nonanoyl-N-methylglucamide and decanoyl-N-methylglucamide were determined to be 25 mM and 7 mM, respectively, by photometric assay. Then solubilization and reconstitution of the melibiose transport carrier were performed using these detergents at concentrations above the CMC. Melibiose counterflow activity was observed with the proteoliposomes reconstituted from the extracted proteins and phospholipids. The proton-translocating ATPase complex (F1-F0) was also solubilized with these detergents. These results indicate that nonanoyl- and decanoyl-N-methylglucamide are useful detergents for solubilization and reconstitution of membrane proteins.

Biological Transport↗
PubMed1984

Temperature dependence of action of polymyxin B on Escherichia coli.

The temperature dependence of the action of polymyxin B on Escherichia coli was studied by using K+, Ca2+, and tetraphenylphosphonium (TPP+) ion-selective electrodes. At room temperature (27 degrees C), Ca2+ was released immediately after addition of polymyxin, while the efflux of K+ occurred after 30 s. The rapid release of Ca2+ was not affected by incubation temperature, while the efflux of K+ was significantly lowered at temperatures below about 25-30 degrees C. The uptake of TPP+ also increased after polymyxin addition. The release of Ca2+ and the uptake of TPP+ supported the disruption of the outer membrane structure reported previously. In experiments with isolated membrane vesicles (the cytoplasmic membrane being exposed), the efflux of K+ was not delayed, but was lowered at temperatures below about 15-20 degrees C. This temperature range differed significantly from that of whole cells, and was interpreted as representing a difference in membrane fluidity between the outer and cytoplasmic membranes. The phase transition temperature of the outer membrane is known to be higher than that of the cytoplasmic membrane; and the temperature dependence of efflux of K+ from membrane vesicles was compatible with the phase transition temperature of liposomes prepared with phospholipids (not containing lipopolysaccharides) extracted from E. coli. Thus, it was speculated that, with whole cells, polymyxin molecules passed through the outer membrane at temperatures above the phase transition and reached the cytoplasmic membrane, increasing its K+ permeability. The mechanism of the permeability change is discussed in terms of deformation of the cytoplasmic membrane structure induced by polymyxin molecules.

Calcium↗
PubMed1984

Use of n-octyl-beta-D-thioglucoside, a new nonionic detergent, for solubilization and reconstitution of membrane proteins.

n-Octyl-beta-D-thioglucopyranoside (octylthioglucoside), a new nonionic detergent, was synthesized. Properties of this detergent and its applicability to membrane biochemistry were investigated. The critical micelle concentration of this detergent was determined to be 9 mM. Membrane proteins of Escherichia coli were effectively solubilized with octylthioglucoside at 25-35 mM. The solubilizing power was equivalent to that of n-octyl-beta-D-glucopyranoside (octylglucoside). Reconstitution of the melibiose carrier into liposomes was performed by the detergent dilution procedure. It was found that the concentration range of octylthioglucoside for successful reconstitution (45-70 mM) was considerably wider than that of octylglucoside (43-46 mM), thus giving more reproducible results. Octylthioglucoside is more stable than octylglucoside. Furthermore, the former is synthesized with high yield at low cost whereas the latter is very expensive. Thus, we conclude that octylthioglucose is superior to octylglucoside, and is very useful in membrane biochemistry.

Bacterial Proteins↗
PubMed1984

Development of polyetherurethane sheet embedded with powdered charcoal for use in hemoperfusion.

This article describes the development of a polyetherurethane sheet embedded with powdered charcoal ( UPC ) for use in a hemoperfusion system. The UPC was 200 micron thick and the release of microparticles could be limited. The powdered charcoal used was selected for optimum pore size distribution and particle size (10-40 micron) to provide a maximum adsorption of bilirubin. The results of an in vitro adsorption test with this material showed an adsorption of bilirubin approximately 14 and 6 times higher than those by bead-type charcoal and XAD-7, respectively. Jaundiced dogs were subjected to direct hemoperfusion through a UPC column containing 36 g powdered charcoal. Effective decreases in bilirubin were observed, giving the maximum removal amounts of 78% in conjugated bilirubin and 76% in unconjugated bilirubin.

Adsorption↗
PubMed1984

Escherichia coli mutants with altered cation recognition by the melibiose carrier.

Revertants that showed normal cation recognition for melibiose transport were isolated from mutants with altered cation recognition (W3133-2S and W3133-2T) of Escherichia coli. Although the original two mutants possessed a second alteration, an increased activity of the Na+(Li+)/H+ antiporter, the revertants, which possessed the normal melibiose carrier, still showed altered properties of the Na+(Li+)/H+ antiporter. These results support the view that the alterations in the melibiose carrier and in the Na+(Li+)/H+ antiporter, observed in the mutants, are not genetically linked.

Escherichia coli↗