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Biomedical subjects

T Ando

Publications and source records attributed to T Ando.

At least 613 records · Page 34Linked to original sources
PubMed1984

Purification of a eukaryotic site-specific endonuclease, Endo.Sce I, from Saccharomyces cerevisiae and effectors on its specificity and activity.

A site-specific endonuclease (Endo.Sce I) which caused double-strand scission of DNA was highly purified from a eukaryote, Saccharomyces cerevisiae IAM4274. The molecular weight of the active form of Endo.Sce I was estimated to be 120,000 and 110,000 by sedimentation analysis on a glycerol density gradient and gel filtration on Ultrogel AcA34, respectively. Analysis of the fractions from the last column chromatography by polyacrylamide gel-electrophoresis in the presence of sodium dodecyl sulfate and by an assay of the endonucleolytic activities suggested that Endo.Sce I consists of two non-identical subunits with molecular weights of 75,000 and 50,000. Unlike restriction endonucleases, Endo.Sce I was active on chromosomal DNA of the cells which produced Endo.Sce I. Single-stranded DNA was not cleaved by Endo.Sce I, but inhibited the endonucleolytic activity of the enzyme on double-stranded DNA. The endonucleolytic activity of Endo.Sce I required the magnesium ions (Mg2+) as a sole cofactor; Mg2+ could not be replaced by Ca2+ or Zn2+. When Mg2+ was replaced by manganese ions (Mn2+), extensively purified Endo.Sce I cleaved double-stranded DNA at many other sites in addition to the sites at which DNA was cleaved in the presence of Mg2+. Experiments indicated that this is not the activation of contaminating endonuclease in the preparation of Endo.Sce I, but the result of relaxation in the site-specificity of cleavage.

Base Sequence↗
PubMed1984

[An electrophysiological method for detecting diabetic retinopathy in rats].

The present study was undertaken to devise an electrophysiological method for detecting diabetic retinopathy in rats. The electroretinogram (ERG) and visual evoked potential (VEP) were recorded from unanesthetized and unrestrained rats rendered diabetic with a single i.v. injection of streptozotocin (STZ) at 35 or 40 mg/kg. The STZ-treated rats showed signs of diabetes: hyperglycemia, glucosuria, hypoinsulinemia, polyuria and increased water intake. Amplitudes of the ERG a- and b-waves and oscillatory potentials (OPs) on the b-wave were decreased and latencies of these waves were prolonged gradually after STZ was administered. Especially, latencies of the OPs became significantly different from the pre-treatment values. Latency of the VEP N1 wave showed a slight prolongation, which might be secondary to the depression of retinal function. Histological examination showed swelling and proliferation of the lens epithelium and swelling and vacuolization of the lens fiber were observed in the eyeball 9 weeks after STZ-treatment. Moreover, thinning of each retinal layer was observed in a few rats. Daily s.c. injection of insulin at 10 units/rat/day started from the 4th week. The ERG values returned to the control values after 2-3 weeks of insulin therapy. These results indicate that the ERG and VEP recording procedure used in the present study is useful for early detection of the diabetic retinopathy in rats and that the OP of the ERG appears to be vulnerable to diabetes in the rat as it is in the human.

Animals↗
PubMed1984

Amauromine, a new vasodilator. Taxonomy, isolation and characterization.

Amauromine is a new alkaloid with vasodilating activity obtained from the culture broth of Amauroascus sp. No. 6237. Its molecular formula was determined to be C32H36N4O2 on the basis of elementary analysis and high resolution mass spectroscopic measurement. It has low toxicity in mice.

Alkaloids↗
PubMed1984

[An ultrastructural and cytochemical study of the rat kidney in experimentally induced endotoxin shock].

The author has investigated the ultrastructural alterations of renal cortex in the shock state which was induced by E.coli endotoxin. The cytochemical study using horseradish peroxidase as a tracer and Na+-K+ ATPase has been performed in an attempt of correlation between morphological changes and functional impairment. In the initial stage after endotoxin administration, vascular changes which might be justified as morphological evidences for microcirculatory derangement were observed. In four hours after administration, aggregates of platelets and fibrin in the glomerular-loop were noted. The proximal tubuli were also involved in various extent of pathological changes compared with the distal tubuli. The increased vascular permeability was substantiated by the peroxidase labeling-method, however, the decreased glomerular permeability was confirmed which may represent the decreased glomerular filtration capacity. The reabsorption of the tracer in the proximal tubuli was initially increased, however, in later, the reabsorption was marked decreased, and similar alteration of membrane-linked Na+-K+ ATPase was also existed in the distal tubuli. In these observations, the tubular reabsorption mechanism was temporarily increased as the rebound reaction to maintain the biological equilibrium, however, on account for persisting microcirculatory derangement, sever functional impairment may occur which eventually leads to irreversible dysfunction of the kidney.

Animals↗
PubMed1984

[Effect of PSK on the recovery of macrophage function in tumor-bearing mice].

Chemotactic response of macrophage to PHA was depressed at early stages in allogeneic sarcoma 180-bearing ICR mice and syngeneic X5563-bearing C3H/He mice and their depressed response were restored to the normal levels by PSK. Macrophages-dependent resistance against Listeria monocytogenes at early phase of infection was depressed at the early stage after tumor inoculation. The depressed resistance was restored by PSK in X5563-bearing mice and by a combination of tumor resection and PSK in sarcoma 180-bearing mice.

Animals↗
PubMed1983

ATP-dependent unwinding of the double helix and extensive supercoiling by Escherichia coli recA protein in the presence of topoisomerase.

recA protein, which is essential for genetic recombination in Escherichia coli, causes extensive unwinding of the double helix by an ATP-dependent reaction and accumulation of positive supercoiling in closed circular double-stranded DNA. Initiation of the extensive unwinding was largely dependent on homologous single-stranded DNA. Therefore, it is likely that the extensive unwinding is initiated mainly at the site of D-loops. "Nascent D-loops" in which the two DNA molecules did not interwind were also good initiation sites of extensive unwinding. When the concentration of Mg2+ was decreased from the standard conditions for D-loop formation (13 mM MgCl2; the higher Mg2+ condition) to the lower Mg2+ condition (1 to 2 mM MgCl2), extensive unwinding by recA protein was initiated very quickly in the absence of single-stranded DNA. Results showed that this single-stranded DNA-independent initiation of extensive unwinding (i) requires negative superhelicity of the double-stranded DNA and (ii) is a first order reaction with respect to the DNA. These observations suggest that, under the lower Mg2+ condition, the extensive unwinding starts at a transiently denatured site in the negative superhelical DNA. Once initiated, the unwinding by recA protein is propagated extensively, even under conditions that do not allow its initiation. Therefore, the propagation of unwinding is a processive reaction ("processive unwinding"). Previous studies indicated that recA protein promotes "distributive unwinding" of double helix which depends on single-stranded DNA. Therefore, recA protein promotes unwinding of the double helix by either of two distinct pathways. Stress caused by the processive unwinding could explain the dissociation of D-loops and reversible inactivation of the double-stranded DNA in a D-loop cycle.

Adenosine Triphosphate↗
PubMed1983

The process in which nucleotide is buried into the active site of heavy meromyosin.

The process in which nucleotide is buried into the active site of heavy meromyosin was studied with stopped-flow apparatus by monitoring the time-course of the large fluorescence increase of 1,N6-ethenoadenosine triphosphate (epsilon-ATP) when it binds from acrylamide-containing solutions. We have recently reported that free epsilon-ATP fluorescence is effectively quenched by acrylamide while bound epsilon-ATP is resistant to quenching by acrylamide. In the present study it was found that in the first step the phosphate moiety binds at a high rate, while the adenine moiety is still on the rim of the active site; the adenine moiety is then pulled into a crevice, and finally epsilon-ATP hydrolysis occurs.

Adenosine Triphosphatases↗
PubMed1983

Correlation of enzyme-induced cleavage sites on negatively superhelical DNA between prokaryotic topoisomerase I and S1 nuclease.

Negatively superhelical pNS1 DNA with a molecular weight of 2.55 MDa (4 kbp) was found to contain 13 specific, unbasepaired sites that are sensitive to a single-strand-specific S1 nuclease cleavage. The S1-cleavage occurred once at these sites. In the absence of added Mg2+, the topoisomerase I purified from Haemophilus gallinarum formed a complex with the superhelical pNS1 DNA which has a hidden strand cleavage. Extensive proteinase K digestion of the complex led to cleavage of the DNA chain. Then the proteinase K-cleaved product was digested with S1, which can cut the opposite strand at the preexisting strand cleavage to generate unit-length linear DNA. Restriction endonuclease analysis of the linear DNA shows that the topoisomerase-induced cleavage occurred once at ten specific sites on the DNA. The topoisomerase caused mainly single-strand cleavage at these sites, but infrequently also caused double-strand cleavage at the same sites. Of interest is the fact that these sites considerably coincide with the S1-cleavable, unbasepaired sites.

Bacillus subtilis↗
PubMed1983

A new class of site-specific endodeoxyribonucleases. Endo.Sce I isolated from a eukaryote, Saccharomyces cerevisiae.

We had found that yeasts had intracellular endodeoxyribonucleases that cut phage DNA into a set of double-stranded fragments with discrete chain lengths. We purified one of them to apparent homogeneity from Saccharomyces cerevisiae and designated it Endo.Sce I. Sequence analysis around 5 cleavage sites in plasmid DNA and phage DNA revealed that Endo.Sce I cuts a defined phosphodiester bond in each strand of double helix at the cleavage sites and produces free cohesive ends consisting of 4 nucleotides protruding at 3'-termini. However, unlike in the case of prokaryotic type II-restriction endonucleases, (i) Endo.Sce I seems to consist of two nonidentical subunits, (ii) no common palindrome or consensus sequence including more than 5 base pairs is detected at or near these cleavage sites, and (iii) Endo.Sce I can cut the DNA isolated from the cells that produced Endo.Sce I. All of the 5 cleavage sites are included in inverted repeats, but these inverted repeats are variable in size, nucleotide sequence, and distance between repeating units. An inverted repeat itself is not a structure recognized by Endo.Sce I. This study shows that Endo.Sce I is the first example of eukaryotic site-specific endonuclease and has properties, as described above, which distinguish it from prokaryotic restriction endonucleases.

Base Sequence↗
PubMed1983

A sensitive spectrophotometric assay for guanase activity.

A highly sensitive and accurate spectrophotometric method was developed for determination of guanase activity with guanine as substrate. The assay is based on the oxidative coupling of 3-methyl-2-benzothiazolinone hydrazone and N,N-diethylaniline. Xanthine formed from guanine by guanase is oxidized to uric acid and hydrogen peroxide by xanthine oxidase, and the hydrogen peroxide produced is determined by an oxidative-coupling reaction with 3-methyl-2-benzothiazolinone hydrazone and N,N-diethylaniline mediated by peroxidase. Formation of the indamine dye is greatly affected by the superoxide radical ion (O2-) and pH value. These problems can be overcome by separating the two reactions of hydrogen peroxide formation and color production and carrying out that color-producing reaction at pH 3.0. This method is very sensitive and accurate because the indamine dye has a very high molar extinction coefficient of 29,800. It can be used with various kinds of automatic analyzers such as a Hitachi, Olympus, or Technicon analyzer. Comparative studies showed that this method is more sensitive and reproducible than other methods. Furthermore, guanase activities determined by this method correlated well with those determined by the improved Ellis-Goldberg method. This method should be useful for measurement of guanase activity in banked blood for preventing transfusion hepatitis and could be valuable as a liver function test.

Aminohydrolases↗
PubMed1983

Specific binding of chartreusin, an antitumor antibiotic, to DNA.

Chartreusin, an antitumor and antibacterial antibiotic, was found to inhibit negatively superhelical DNA-relaxation catalyzed by prokaryotic topoisomerase I and conversion of the superhelical DNA into unit length linear form catalyzed by single-strand-specific S1 nuclease. The inhibitory effect of the agent was due to the binding to DNA causing the alteration of tertiary structure. To characterize the binding specificity, we investigated the protection of DNA against cleavages by various restriction endonucleases. It was evidenced that the binding of the agent is not at random and correlates to the sequence 5'CGC 3' 3'GCG 5' on DNA stretch.

Antibiotics, Antineoplastic↗