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61.
Hiroyuki Morimoto Hirohiko Okamura Kaya Yoshida Seiichiro Kitamura Tatsuji Haneji 《Journal of enzyme inhibition and medicinal chemistry》2013,28(4):327-331
The reversible phosphorylation of proteins mediates cellular signals in eukaryotic cells. RNA interference inhibits the expression of genes and proteins in a sequence-specific manner and provides a tool to study the functions of target molecules. The effect of RNA interference on protein phosphatase isoforms in HEK-293 cells was examined. Protein phosphatase 1 delta (PP1δ) sequence-specific double-stranded RNA (dsRNA) inhibited mRNA and protein expression of the PP18. This RNA interference did not affect the expression of α and γ1 isoforms of PP1. Transfection of antisense RNA specific for PP1δ also suppressed the expression of PP1δ. It was further demonstrated by an in vitro RNA cleavage assay that extracts of HEK-293 cells catalyzed the processing of dsRNA. This cell line had much stronger mRNA expression of Dicer, an RNase III-like enzyme, than did human osteoblastic MG63 cells. The present results show that RNA interference is a useful tool to distinguish between PP1 isoforms. 相似文献
62.
Mitsuo. Sekine Hiroyuki. Tsuruoka Koh-ichiro. Shohda Tomohisa. Moriguchi Tomohisa. Wada 《Nucleosides, nucleotides & nucleic acids》2013,32(9-11):2033-2043
Abstract This paper describes general methods for the synthesis of N-phosphorylated ribonucleosides and oligonucleotides containing a 2′-O-phosphorylated or 2′-O-thiophosphorylated ribonucleoside. The NMR-based conformational analysis and computational molecular dynamics simulation of the 2′-O-phosphorylated ribonucleoside residue in such modified oligonucleotides suggested that the ribose residue existed preferentially in a C2′-endo conformation. It was also found that simple heating of 2′-O-phosphorylated oligonucleotides resulted in rapid dethiophosphorylation. 相似文献
63.
Koko Moriya Kei Nagatoshi Yoshimi Noriyasu Tsuyoshi Okamura Emi Takamitsu Takashi Suzuki Toshihiko Utsumi 《PloS one》2013,8(11)
N-myristoylation of eukaryotic cellular proteins has been recognized as a modification that occurs mainly on cytoplasmic proteins. In this study, we examined the membrane localization, membrane integration, and intracellular localization of four recently identified human N-myristoylated proteins with predicted transmembrane domains. As a result, it was found that protein Lunapark, the human ortholog of yeast protein Lnp1p that has recently been found to be involved in network formation of the endoplasmic reticulum (ER), is an N-myristoylated polytopic integral membrane protein. Analysis of tumor necrosis factor-fusion proteins with each of the two putative transmembrane domains and their flanking regions of protein Lunapark revealed that transmembrane domain 1 and 2 functioned as type II signal anchor sequence and stop transfer sequence, respectively, and together generated a double-spanning integral membrane protein with an N-/C-terminal cytoplasmic orientation. Immunofluorescence staining of HEK293T cells transfected with a cDNA encoding protein Lunapark tagged with FLAG-tag at its C-terminus revealed that overexpressed protein Lunapark localized mainly to the peripheral ER and induced the formation of large polygonal tubular structures. Morphological changes in the ER induced by overexpressed protein Lunapark were significantly inhibited by the inhibition of protein N-myristoylation by means of replacing Gly2 with Ala. These results indicated that protein N-myristoylation plays a critical role in the ER morphological change induced by overexpression of protein Lunapark. 相似文献
64.
Takao Tanahashi Tomohisa Yamamoto Takuyuki Endo Harutoshi Fujimura Masaru Yokoe Hideki Mochizuki Taishin Nomura Saburo Sakoda 《PloS one》2013,8(12)
Freezing of gait in patients with Parkinson’s disease is associated with several factors, including interlimb incoordination and impaired gait cycle regulation. Gait analysis in patients with Parkinson’s disease is confounded by parkinsonian symptoms such as rigidity. To understand the mechanisms underlying freezing of gait, we compared gait patterns during straight walking between 9 patients with freezing of gait but little to no parkinsonism (freezing patients) and 11 patients with Parkinson’s disease (non-freezing patients). Wireless sensors were used to detect foot contact and toe-off events, and the step phase of each foot contact was calculated by defining one stride cycle of the other leg as 360°. Phase-resetting analysis was performed, whereby the relation between the step phase of one leg and the subsequent phase change in the following step of the other leg was quantified using regression analysis. A small slope of the regression line indicates a forceful correction (phase reset) at every step of the deviation of step phase from the equilibrium phase, usually at around 180°. The slope of this relation was smaller in freezing patients than in non-freezing patients, but the slope exhibited larger step-to-step variability. This indicates that freezing patients executed a forceful but noisy correction of the deviation of step phase, whereas non-freezing patients made a gradual correction of the deviation. Moreover, freezing patients tended to show more variable step phase and stride time than non-freezing patients. Dynamics of a model of two coupled oscillators interacting through a phase resetting mechanism were examined, and indicated that the deterioration of phase reset by noise provoked variability in step phase and stride time. That is, interlimb coordination can affect regulation of the gait cycle. These results suggest that noisy interlimb coordination, which probably caused forceful corrections of step phase deviation, can be a cause of freezing of gait. 相似文献
65.
Kentaro Inoue Jun Wada Jun Eguchi Atsuko Nakatsuka Sanae Teshigawara Kazutoshi Murakami Daisuke Ogawa Takahiro Terami Akihiro Katayama Atsuhito Tone Izumi Iseda Kazuyuki Hida Masao Yamada Tomohisa Ogawa Hirofumi Makino 《PloS one》2013,8(10)
We analyzed the urine samples of patients with type 2 diabetes at various stages of diabetic nephropathy by lectin microarray to identify a biomarker to predict the progression of diabetic nephropathy. Japanese patients with type 2 diabetes at various stages of nephropathy were enrolled and we performed lectin microarray analyses (n = 17) and measured urinary excretion of fetuin-A (n = 85). The increased signals of urine samples were observed in Siaα2-6Gal/GalNAc-binding lectins (SNA, SSA, TJA-I) during the progression of diabetic nephropathy. We next isolated sialylated glycoproteins by using SSA-lectin affinity chromatography and identified fetuin-A by liquid chromatography–tandem mass spectrometer. Urinary excretion of fetuin-A significantly increased during the progression of albuminuria (A1, 0.40±0.43; A2, 0.60±0.53; A3 1.57±1.13 ng/gCr; p = 7.29×10−8) and of GFR stages (G1, 0.39±0.39; G2, 0.49±0.45; G3, 1.25±1.18; G4, 1.34±0.80 ng/gCr; p = 3.89×10−4). Multivariate logistic regression analysis was employed to assess fetuin-A as a risk for diabetic nephropathy with microalbuminuria or GFR<60 mL/min. Fetuin-A is demonstrated as a risk factor for both microalbuminuria and reduction of GFR in diabetic nephropathy with the odds ratio of 4.721 (1.881–11.844) and 3.739 (1.785–7.841), respectively. Collectively, the glycan profiling analysis is useful method to identify the urine biomarkers and fetuin-A is a candidate to predict the progression of diabetic nephropathy. 相似文献
66.
Hiroaki Tobimatsu Antoine Paragon Yosuke Okamura Shinji Takeoka Ryo Sudo Yasuo Ikeda Kazuo Tanishita 《Journal of Biorheology》2013,26(1-2):11-20
Recombinant glycoprotein Ibα latex beads (rGPIbα-LB) are a potential solution to overcoming platelet transfusion problems with artificial platelets. To understand the transport process of artificial platelets and to estimate the particle motion when adhering to the wall surface, we evaluated the lateral motion of rGPIbα-LB in terms of drift and random motion, because the lateral motion is an important factor for transport and adhesion. We observed the lateral motion of rGPIbα-LB flowing with red blood cells toward the immobilized von Willebrand factor (vWf) surface in a model arteriole at wall shear rates of 200–1000 s?1 and 0–40% Hct. At 40% Hct, wall shear rate dependence was observed for the drift motion, i.e. the lateral velocity of rGPIbα-LB toward the wall. In the near-wall region, the drift motion of contacting particles differed substantially from that of non-contacting particles. Additionally, the trajectories of contacting particles on the vWf surface had specific motion that was not observed on the BSA surface. These results suggest that the adhesion force between rGPIbα and vWf is highly associated with the motion of particles near the wall. These features are desirable for artificial platelets, particularly for the adhesion process. 相似文献
67.
Yumi Mori Takafumi Kataoka Takahiko Okamura Ryuji Kondo 《Archives of microbiology》2013,195(5):303-312
This study investigated the spatiotemporal abundance and diversity of the α-subunit of the dissimilatory sulfite reductase gene (dsrA) in the meromictic Lake Suigetsu for assessing the sulfur-oxidizing bacterial community. The density of dsrA in the chemocline reached up to 3.1 × 106 copies ml?1 in summer by means of quantitative real-time PCR and it was generally higher than deeper layers. Most of the dsrA clones sequenced were related to green sulfur bacteria such as Chlorobium phaeovibrioides, C. limicola, and C. luteolum. Below the chemocline of the lake, we also detected other dsrA clones related to the purple sulfur bacterium Halochromatium salexigens and some branching lineages of diverse sequences that were related to chemotrophic sulfur bacterial species such as Magnetospirillum gryphiswaldense, Candidatus Ruthia magnifica, and Candidatus Thiobios zoothamnicoli. The abundance and community compositions of sulfur-oxidizing bacteria changed depending on the water depth and season. This study indicated that the green sulfur bacteria dominated among sulfur-oxidizing bacterial population in the chemocline of Lake Suigetsu and that certain abiotic environmental variables were important factors that determined sulfur bacterial abundance and community structure. 相似文献
68.
69.
The voltage-sensor domain (VSD) is a functional module that undergoes structural transitions in response to membrane potential changes and regulates its effectors, thereby playing a crucial role in amplifying and decoding membrane electrical signals. Ion-conductive pore and phosphoinositide phosphatase are the downstream effectors of voltage-gated channels and the voltage-sensing phosphatase, respectively. It is known that upon transition, the VSD generally acts on the region C-terminal to S4. However, whether the VSD also induces any structural changes in the N-terminal region of S1 has not been addressed directly. Here, we report the existence of such an N-terminal effect. We used two distinct optical reporters—one based on the Förster resonance energy transfer between a pair of fluorescent proteins, and the other based on fluorophore-labeled HaloTag—and studied the behavior of these reporters placed at the N-terminal end of the monomeric VSD derived from voltage-sensing phosphatase. We found that both of these reporters were affected by the VSD transition, generating voltage-dependent fluorescence readouts. We also observed that whereas the voltage dependencies of the N- and C-terminal effects appear to be tightly coupled, the local structural rearrangements reflect the way in which the VSD is loaded, demonstrating the flexible nature of the VSD. 相似文献
70.
Pseudomonas M16 is the mutant derived from a facultative methylotroph, Pseudomonas N842, which is the potent producer of coenzyme Q10 (CoQ10). This mutant with elevated productivity of CoQ10 was observed to accumulate the significant amount of another CoQ homolog, which could not be detected in the parent strain. This CoQ homolog was extracted from the intact cells of the mutant and purified to crystaline state. The chemical properties and the results of UV, NMR and mass spectrometries revealed that this CoQ homolog was CoQ11. 相似文献