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81.
Daisuke Harada Shinsaku Naito Yoshiyuki Kawauchi Keiko Ishikawa Osamu Koshitani Isao Hiraoka Masaki Otagiri 《Analytical biochemistry》2001,290(2)
A high-performance liquid chromatographic assay was developed for the quantitative determination of the sulfur-containing amino acids N-acetyl-
-cysteine (NAC) and
-cysteine (Cys) in rat plasma. The thiols were separated by reverse-phase ion-pair chromatography, and the column eluent was continuously mixed with an iodoplatinate-containing solution. The substitution of sulfur of the thiol compound with iodide was quantitatively determined by measuring changes in the absorption at 500 nm. The low-molecular-weight disulfides and mixed disulfide conjugates of thiols with proteins were entirely reduced to the original reduced compounds by dithiothreitol. By reducing these two types of disulfides separately during sample pretreatment, the reduced, protein-unbound, and total thiol concentrations could also be determined. Validation testing was performed, and no problems were encountered. The limit of detection was approximately 20 pmol of thiol on the column. The present method was used to measure the plasma concentrations of NAC and Cys in the rat after a bolus intravenous administration of NAC, focusing on disulfide formation. The binding of NAC to protein through mixed disulfide formation proceeds in a time-dependent and reversible manner. Moreover, this “stable” covalent binding might limit total drug elimination, while the unbound NAC is rapidly eliminated. Consequently, the analytical method described in this study is very useful for the determination of plasma NAC and Cys, including disulfide conjugates derived from them. 相似文献
82.
Dynamic Behavior of Microtubules during Dynein-dependent Nuclear Migrations of Meiotic Prophase in Fission Yeast 总被引:1,自引:0,他引:1 下载免费PDF全文
Ayumu Yamamoto Chihiro Tsutsumi Hiroaki Kojima Kazuhiro Oiwa Yasushi Hiraoka 《Molecular biology of the cell》2001,12(12):3933-3946
During meiotic prophase in fission yeast, the nucleus migrates back and forth between the two ends of the cell, led by the spindle pole body (SPB). This nuclear oscillation is dependent on astral microtubules radiating from the SPB and a microtubule motor, cytoplasmic dynein. Here we have examined the dynamic behavior of astral microtubules labeled with the green fluorescent protein during meiotic prophase with the use of optical sectioning microscopy. During nuclear migrations, the SPB mostly follows the microtubules that extend toward the cell cortex. SPB migrations start when these microtubules interact with the cortex and stop when they disappear, suggesting that these microtubules drive nuclear migrations. The microtubules that are followed by the SPB often slide along the cortex and are shortened by disassembly at their ends proximal to the cortex. In dynein-mutant cells, where nuclear oscillations are absent, the SPB never migrates by following microtubules, and microtubule assembly/disassembly dynamics is significantly altered. Based on these observations, together with the frequent accumulation of dynein at a cortical site where the directing microtubules interact, we propose a model in which dynein drives nuclear oscillation by mediating cortical microtubule interactions and regulating the dynamics of microtubule disassembly at the cortex. 相似文献
83.
Y Wataya H Satake O Hiraoka T Aji K Morishige J Y Kimura A Ishii A Matsuda T Ueda K Fukukawa 《Nucleic acids symposium series》1986,(17):149-151
Carbocyclic inosine is a potent inhibitor for the growth of the promastigote form of Leishmania tropica and Leishmania donovani. In culture, the EC50 values of carbocyclic inosine are 8.3 X 10(-8) and 1.3 X 10(-7) M for the promastigotes of L. tropica and L. donovani, respectively. On the other hand, it is less toxic towards mouse mammary tumor FM3A cells: the EC50 value is 2.7 X 10(-4)M. Carbocyclic inosine is metabolized by Leishmania promastigotes to give carbocyclic adenosine-5'-triphosphate(aristeromycin-5'-triphosphate) and carbocyclic guanosine-5'-triphosphate. This metabolic conversion provides a mechanism for the parasite-selective toxicity of carbocyclic inosine. Carbocyclic inosine was found to be active against L. donovani amastigotes in an in vivo-like cultivation in vitro. 相似文献
84.
85.
86.
Immunochemical properties and intracellular localization of two molecular forms of arginine aminopeptidase in Streptococcus mitis ATCC 9811 总被引:1,自引:0,他引:1
B Y Hiraoka 《Biochimica et biophysica acta》1985,841(2):166-172
Streptococcus mitis contains two multiple forms of arginine aminopeptidase (I and II) which differ from each other with respect to their content, immunochemical properties and cellular localization. Immunological analyses by Ouchterlony double immunodiffusion and immunoprecipitation showed an antigenic difference between each form by the use of antisera specific for each enzyme. The amounts of enzymes I and II within the cell were estimated to be 230 +/- 4.3 and 646 +/- 20 ng/mg protein (+/- S.D.), respectively, using a standard curve of purified enzyme in a single radial immunodiffusion assay. When intact cells were treated with the cell wall lytic enzyme, N-acetylmuramidase, though both enzymes were solubilized, a time lag was observed for the solubilization of enzyme II. Enzyme I was detected only in the cell wall fraction and showed no detectable associated with the membrane. Although most of the enzyme II activity was recovered in the cell wall fraction, a slight amount (7.5%) of the total activity was also found in the membrane fraction. 相似文献
87.
88.
Electrophysiological properties of isolated rat liver cells 总被引:2,自引:0,他引:2
T Sawanobori H Takanashi M Hiraoka Y Iida K Kamisaka H Maezawa 《Journal of cellular physiology》1989,139(3):580-585
The electrophysiological properties of isolated rat liver cells were studied using the patch clamp method in whole-cell configuration. The membrane potential in isolated hepatocytes was -42 +/- 7 mV (n = 20). The input resistance (Rin) and the time constant (tau m) were 51 +/- 17 M (the range of 34 to 180 M omega) (n = 20) and 4.2 +/- 1.0 msec (the range of 3 to 16.5 ms) (n = 20). Assuming that the specific membrane capacitance is 1 microF/cm2, the membrane resistance and membrane capacitance were 42. +/- 9.0 K omega cm2 and 87 +/- 27 pF. These values indicate that isolated rat hepatocytes are not abnormally permeable or leaky. The current-voltage relationship was linear with no rectification. The depolarizing pulse from the resting potential did not induce fast or slow inward currents even when norepinephrine or high Ca2 (3.6 mM) were applied. This indicates that there is no voltage-sensitive Ca2+ channel in the isolated hepatocytes. 相似文献
89.
Tazro Ohta Takeshi Kawashima Natsuko O. Shinozaki Akito Dobashi Satoshi Hiraoka Tatsuhiko Hoshino Keiichi Kanno Takafumi Kataoka Shuichi Kawashima Motomu Matsui Wataru Nemoto Suguru Nishijima Natsuki Suganuma Haruo Suzuki Y-h. Taguchi Yoichi Takenaka Yosuke Tanigawa Momoka Tsuneyoshi Kazutoshi Yoshitake Yukuto Sato Riu Yamashita Kazuharu Arakawa Wataru Iwasaki 《Journal of plant research》2018,131(4):709-717
Recent studies have shown that environmental DNA is found almost everywhere. Flower petal surfaces are an attractive tissue to use for investigation of the dispersal of environmental DNA in nature as they are isolated from the external environment until the bud opens and only then can the petal surface accumulate environmental DNA. Here, we performed a crowdsourced experiment, the “Ohanami Project”, to obtain environmental DNA samples from petal surfaces of Cerasus?×?yedoensis ‘Somei-yoshino’ across the Japanese archipelago during spring 2015. C. × yedoensis is the most popular garden cherry species in Japan and clones of this cultivar bloom simultaneously every spring. Data collection spanned almost every prefecture and totaled 577 DNA samples from 149 collaborators. Preliminary amplicon-sequencing analysis showed the rapid attachment of environmental DNA onto the petal surfaces. Notably, we found DNA of other common plant species in samples obtained from a wide distribution; this DNA likely originated from the pollen of the Japanese cedar. Our analysis supports our belief that petal surfaces after blossoming are a promising target to reveal the dynamics of environmental DNA in nature. The success of our experiment also shows that crowdsourced environmental DNA analyses have considerable value in ecological studies. 相似文献