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101.
102.
Asai R Ootani K Nomura Y Nakamura C Ikebukuro K Arikawa Y Miyake J Karube I 《Marine biotechnology (New York, N.Y.)》2003,5(5):417-423
A technique for detecting Raphidophycean, a bloom-forming genus of algae, was developed using a
specific DNA probe. The design of the probe was based on a sequence polymorphism within the small subunit
(SSU) ribosomal RNA gene (rDNA) of this strain by using fluorescence polarization (FP) analysis and the
BIAcore 2000 biosensor, which utilized surface plasmon resonance (SPR). The specific sequence in SSU rDNA
for Heterosigma carterae was determined by sequence data analysis. One pair of polymerase chain reaction
(PCR) probes was designed for use in making the identification. H. carterae SSU rDNA was amplified by PCR.
Using a fluoroscein isothiocyanate–labeled or biotin-labeled oligonucleotide probe, the PCR-amplified rDNA
was selectively detected as an FP-intensity change via FP analysis or as a resonance-unit change via SPR.
Although total time for final detection after sampling was within 3 hours, specific rDNA could be detected
within 10 minutes after PCR through these detection methods. 相似文献
103.
Association of human origin recognition complex 1 with chromatin DNA and nuclease-resistant nuclear structures 总被引:5,自引:0,他引:5
Tatsumi Y Tsurimoto T Shirahige K Yoshikawa H Obuse C 《The Journal of biological chemistry》2000,275(8):5904-5910
An origin recognition complex (ORC) consisting of six polypeptides has been identified as a DNA replication origin-binding factor in Saccharomyces cerevisiae. Homologues of ORC subunits have been discovered among eukaryotes, and we have prepared monoclonal antibodies against a human homologue of ORC1 (hORC1) to study its localization in human cells. It was thus found to associate with nuclei throughout the cell cycle and to be resistant to nonionic detergent treatment, in contrast to MCM proteins, which are other replication factors, the association of which with nuclei is clearly dependent on the phase of the cell cycle. A characteristic feature of hORC1 is dissociation by NaCl in a narrow concentration range around 0.25 M, suggesting interaction with some specific partner(s) in nuclei. Nuclease treatment experiments and UV cross-linking experiments further indicated interaction with both nuclease-resistant nuclear structures and chromatin DNA. Although its DNA binding was unaffected, some variation in the cell cycle was apparent, the association with nuclear structures being less stable in the M phase. Interestingly, the less stable association occurred concomitantly with hyperphosphorylation of hORC1, suggesting that this hyperphosphorylation may be involved in M phase changes. 相似文献
104.
Motomu Hashimoto Toru Yamazaki Masahide Hamaguchi Takeshi Morimoto Masashi Yamori Keita Asai Yu Isobe Moritoshi Furu Hiromu Ito Takao Fujii Chikashi Terao Masato Mori Takashi Matsuo Hiroyuki Yoshitomi Keiichi Yamamoto Wataru Yamamoto Kazuhisa Bessho Tsuneyo Mimori 《PloS one》2015,10(4)
Purpose
To determine whether the presence of periodontitis (PD) and Porphyromonas gingivalis (Pg) in the subgingival biofilm associates with the development of rheumatoid arthritis (RA) in treatment naïve preclinical stage of arthritis patients.Methods
We conducted a prospective cohort study of 72 consecutive patients with arthralgia who had never been treated with any anti-rheumatic drugs or glucocorticoids. Periodontal status at baseline was assessed by dentists. PD was defined stringently by the maximal probing depth≧4 mm, or by the classification by the 5th European Workshop in Periodontology (EWP) in 2005 using attachment loss. Up to eight plaque samples were obtained from each patient and the presence of Pg was determined by Taqman PCR. The patients were followed up for 2 years and introduction rate of methotrexate (MTX) treatment on the diagnosis of RA was compared in patients with or without PD or Pg.Results
Patients with PD (probing depth≧4mm) had higher arthritis activity (p = 0.02) and higher risk for future introduction of MTX treatment on the diagnosis of RA during the follow up than patients without PD (Hazard ratio 2.68, p = 0.03). Arthritis activity and risk for MTX introduction increased with the severity of PD assessed by EWP, although not statistically significant. On the other hand, presence of Pg was not associated with arthritis activity (p = 0.72) or the risk for MTX introduction (p = 0.45).Conclusion
In treatment naïve arthralgia patients, PD, but not the presence of Pg, associates with arthritis activity and future requirement of MTX treatment on the diagnosis of RA. 相似文献105.
Hiroko Fukushima Takashi Fukushima Aiko Sakai Ryoko Suzuki Chie Kobayashi Yoshiko Oshiro Masashi Mizumoto Noriko Hoshino Chikashi Gotoh Yasuhisa Urita Hiroaki Komuro Michio Kaneko Noritoshi Sekido Kouji Masumoto Hideyuki Sakurai Ryo Sumazaki 《Reports of Practical Oncology and Radiotherapy》2015,20(3):217-222
Background
Rhabdomyosarcoma (RMS) is one of the most common soft tissue sarcomas among children. Patients who developed genitourinary/pelvic rhabdomyosarcoma (GU/P-RMS) have a higher complication ratio and relatively poorer event free survival, with local therapy being very important. While proton beam therapy (PBT) is expected to reduce co-morbidity, especially for children, this lacks firm evidence and analysis. We analyzed GU/P-RMS children who had undergone multimodal therapy combined with PBT at a single institution.Method
We retrospectively reviewed charts of children with GU/P-RMS treated from January 2007 to May 2013 at the University of Tsukuba Hospital who had undergone multimodal therapy with PBT.Results
There were 5 children and their median age at diagnosis was 2.8 years (0.6–4.4 years). Primary sites were the bladder (2) and the prostate (3). All received neo-adjuvant chemotherapy and 3 underwent chemotherapy during PBT (Group Cx). All patients of Group Cx developed leukocytopenia (WBC <1000/μL). The median dose of PBT was 47.7 GyE (41.4–50.4 GyE). All patients survived by their last hospital visit (median, 36 months).Conclusions
We analyzed multimodal treatment combined with PBT applied for GU/P-RMS. PBT was well tolerated and could be a plausible choice instead of photon therapy for this population. 相似文献106.
Takanori Kihara Narutoshi Yoshida Shingo Mieda Kyoko Fukazawa Chikashi Nakamura Kazuhiko Ishihara Jun Miyake 《NanoBioTechnology》2007,3(2):127-134
Previously, we developed a new molecular delivery system to target single living cells by using atomic force microscope and
ultrathin needle referred to as nanoneedle. This system delivers molecules into the cell by attaching them to the surface
of nanoneedle. However, nonspecific protein adsorption on the nanoneedle surface inside the living cells limits the range
of application of this system. In the present study, we focused on nonspecific protein adsorption onto the nanoneedle surface
inside the cells and examined whether this protein adsorption was reduced by modifying the nanoneedle surface with a biocompatible
phospholipid polymer containing 2-methacryloyloxyethyl phosphorylcholine (MPC) unit. MPC polymer coating of the surface of
silicon wafer reduced nonspecific adsorption of proteins from liver extracts and prevented the formation of clot-like protein
aggregates. MPC polymer also decreased nonspecific adsorption of cytosolic protein onto the nanoneedle surface inside the
living cell. On the other hand, MPC polymer showed no effect on nonspecific mechanical interaction between nanoneedle and
the cell components. Surface modification with MPC polymer is a useful technique to modify the surface properties of nanoneedle. 相似文献
107.
A novel sporulation-deficient mutant, sev4-L5, was isolated in a genetic screen of a collection of temperature-sensitive mutants of Schizosaccharomyces pombe. The wild-type sev4 gene was identified as cta4+, which encodes a putative cation-transporting P-type ATPase. The sev4-L5 allele harbored a single missense mutation that caused replacement of Gly615 with a glutamate at the putative ATP-binding site. Similar to cta4-null mutants, sev4-L5 exhibited defects in growth at high and low temperatures, and sensitivity to high and extremely low concentrations of Ca2+. The cta4+ mRNA level was considerably enhanced during meiosis. When sev4-L5 cells were incubated in sporulation medium at the permissive temperature, meiotic nuclear divisions proceeded with normal kinetics, but spores were not formed. Structural alteration of the spindle pole body, which is prerequisite to construction of the forespore membrane in wild type, was incomplete. Consequently, formation of the forespore membrane was severely impaired. These observations show that perturbation of Ca2+ homeostasis by mutation of cta4/sev4 blocks sporulation mainly by interfering with forespore membrane assembly. 相似文献
108.
Crystalline forms of detergent-solubilized sarcoplasmic reticulum Ca2+-ATPase, obtained in the presence of either a substrate analog, AMPPCP, or a transition state complex, ADP.fluoroaluminate, were recently described to share the same general architecture despite the fact that, when studied in a test tube, these forms show different functional properties. Here, we show that the differences in the properties of the E1.AMPPCP and the E1.ADP.AlFx membraneous (or solubilized) forms are much less pronounced when these properties are examined in the presence of 10 mM Ca2+ (the concentration prevailing in the crystallization media) than when they are examined in the presence of the few micromolar of Ca2+ known to be sufficient to saturate the transport sites. This concerns various properties, including ATPase susceptibility to proteolytic cleavage by proteinase K, ATPase reactivity toward SH-directed Ellman's reagent, ATPase intrinsic fluorescence properties (here described for the E1.ADP.AlFx complex for the first time), and also the rates of 45Ca2+-40Ca2+ exchange at site "II." These results solve the above paradox at least partially and suggest that the presence of a previously unrecognized Ca2+ ion in the E1.AMPPCP crystals should be re-investigated. A contrario, they emphasize the fact that the average conformation of the E1.AMPPCP complex under usual conditions in the test tube differs from that found in the crystalline form. The extended conformation of nucleotide revealed by the E1.AMPPCP crystalline form might be only indicative of the requirements for further processing of the complex, toward the transition state leading to phosphorylation and Ca2+ occlusion. 相似文献
109.
Xu C Ma H Inesi G Al-Shawi MK Toyoshima C 《The Journal of biological chemistry》2004,279(17):17973-17979
Mutational analysis of amino acid residues lining the thapsigargin (TG) binding cavity at the interface of the membrane surface and cytosolic headpiece was performed in the Ca(2+) ATPase (SERCA-1). Specific mutations such as F256V, I765A, and Y837A reduce not only the apparent affinity of the ATPase for TG but also the maximal inhibitory effect. The effect of mutations is dependent on the type and size of the substitute side chain, indicating that hydrophobic partitioning of TG and complementary molecular shapes are involved not only in binding but also in the inhibitory mechanism. A major factor determining the inhibitory effect of bound TG is its interference with conformational changes that are required for the progress of the ATPase cycle. Most prominent and specific is the TG interference with a wide displacement of the Phe-256 side chain that is associated with the E2 to E1.2Ca(2+) transition. The specificity of the TG inhibitory mechanism is emphasized by the finding that the F256V mutation does not interfere at all with the effect of 2,5-di-(t-butyl)-hydroquinone, which is another SERCA inhibitor bound by hydrophobic partitioning. The specificity of the inhibitory mechanism is also emphasized by the observation that within the concentration range producing total inhibition of wild-type SERCA-1, TG produces a 4-fold stimulation of the P-glycoprotein (multidrug transporter) ATPase. 相似文献
110.