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排序方式: 共有535条查询结果,搜索用时 15 毫秒
491.
Putnam CD Clancy SB Tsuruta H Gonzalez S Wetmur JG Tainer JA 《Journal of molecular biology》2001,311(2):297-310
The RuvB hexamer is the chemomechanical motor of the RuvAB complex that migrates Holliday junction branch-points in DNA recombination and the rescue of stalled DNA replication forks. The 1.6 A crystal structure of Thermotoga maritima RuvB together with five mutant structures reveal that RuvB is an ATPase-associated with diverse cellular activities (AAA+-class ATPase) with a winged-helix DNA-binding domain. The RuvB-ADP complex structure and mutagenesis suggest how AAA+-class ATPases couple nucleotide binding and hydrolysis to interdomain conformational changes and asymmetry within the RuvB hexamer implied by the crystallographic packing and small-angle X-ray scattering in solution. ATP-driven domain motion is positioned to move double-stranded DNA through the hexamer and drive conformational changes between subunits by altering the complementary hydrophilic protein- protein interfaces. Structural and biochemical analysis of five motifs in the protein suggest that ATP binding is a strained conformation recognized both by sensors and the Walker motifs and that intersubunit activation occurs by an arginine finger motif reminiscent of the GTPase-activating proteins. Taken together, these results provide insights into how RuvB functions as a motor for branch migration of Holliday junctions. 相似文献
492.
Lata R Conway JF Cheng N Duda RL Hendrix RW Wikoff WR Johnson JE Tsuruta H Steven AC 《Cell》2000,100(2):253-263
Typical of DNA bacteriophages and herpesviruses, HK97 assembles in two stages: polymerization and maturation. First, capsid protein polymerizes into closed shells; then, these precursors mature into larger, stabler particles. Maturation is initiated by proteolysis, producing a metastable particle primed for expansion-the major structural transition. We induced expansion in vitro by acidic pH and monitored the resulting changes by time-resolved X-ray diffraction and cryo-electron microscopy. The transition, which is not synchronized over the population, proceeds in a series of stochastically triggered subtransitions. Three distinct intermediates were identified, which are comparable to transitional states in protein folding. The intermediates' structures reveal the molecular events occurring during expansion. Integrated into a movie (see Dynamic Visualization below), they show capsid maturation as a dynamic process. 相似文献
493.
Yoshida R Yoshioka T Yamane S Matsutani T Toyosaki-Maeda T Tsuruta Y Suzuki R 《Immunogenetics》2000,52(1-2):35-45
We developed an adaptor ligation PCR-based microplate hybridization assay (MHA) to analyze the repertoires of mouse T-cell receptor (TCR) alpha- and beta-chain variable regions (TCRAV and TCRBV). RNA is transcribed to cDNA and an adaptor is ligated to the 5' end of the cDNA, which is then used as a template for PCR with an adaptor-specific 3' primer and a constant region-specific 5' primer. After hybridization of PCR products with TCRAV-and TCRBV-specific probes on the microplate, quantitative ELISA was carried out.The entire TCRAV or TCRBV repertoires could be analyzed using a single 96-well plate in triplicate and completed in less than 4 h. The assay results demonstrated the high level of specificity and reproducibility of this method. Furthermore, MHA results correlated well with those of fluorescence-activated cell sorting. This method may provide important information about various T-cell-associated diseases including autoimmune disease. The influence of the MHC on mouse TCR repertoires was next studied using the newly developed mouse TCRAV and TCRBV repertoire assay. The analysis in six strains showed no significant correlation between MHC haplotypes and TCRAV and TCRBV repertoires. However, large differences among strains was observed in TCRBV, but not in TCRAV repertoires. There were also large differences within same strain in TCRBV, but not in TCRAV repertoires, indicating differences in individuals independent of genetic factors. These data suggest that TCRBV repertoires are more susceptible than TCRAV repertoires not only to genetic factors but also some environmental factors. 相似文献
494.
495.
Alberti MO Roth JC Ismail M Tsuruta Y Abraham E Pereboeva L Gerson SL Curiel DT 《PloS one》2012,7(5):e37812
The gene therapy field is currently limited by the lack of vehicles that permit efficient gene delivery to specific cell or tissue subsets. Native viral vector tropisms offer a powerful platform for transgene delivery but remain nonspecific, requiring elevated viral doses to achieve efficacy. In order to improve upon these strategies, our group has focused on genetically engineering targeting domains into viral capsid proteins, particularly those based on adenovirus serotype 5 (Ad5). Our primary strategy is based on deletion of the fiber knob domain, to eliminate broad tissue specificity through the human coxsackie-and-adenovirus receptor (hCAR), with seamless incorporation of ligands to re-direct Ad tropism to cell types that express the cognate receptors. Previously, our group and others have demonstrated successful implementation of this strategy in order to specifically target Ad to a number of surface molecules expressed on immortalized cell lines. Here, we utilized phage biopanning to identify a myeloid cell-binding peptide (MBP), with the sequence WTLDRGY, and demonstrated that MBP can be successfully incorporated into a knob-deleted Ad5. The resulting virus, Ad.MBP, results in specific binding to primary myeloid cell types, as well as significantly higher transduction of these target populations ex vivo, compared to unmodified Ad5. These data are the first step in demonstrating Ad targeting to cell types associated with inflammatory disease. 相似文献
496.
Proper regulation of morphological changes in neuronal cells is essential for their differentiation. Complex signaling mechanisms mediate a variety of morphological changes such as formation of neurites. It is well established that a number of small GTPases control neurite behavior before the connection with the target tissue. However, their regulatory mechanisms remain to be fully understood. Here, we show that the Arf6 guanine-nucleotide exchange factor (GEF), cytohesin-2 (CYTH2), interacts with the cytoskeletal protein actinin-1 (ACTN1) and regulates neurite extension in N1E-115 cells used as the model. Knockdown of ACTN1, as well as that of CYTH2, in cells inhibits cellular Arf6 activity and neurite extension. The C-terminal polybasic region of CYTH2 participates in interacting directly with the EFh2 domain of ACTN1. Expression of CYTH2 mutant deficient of the EFh2 domain in cells also inhibits Arf6 activation and neurite extension. Furthermore, FRET analysis detects that the respective interactive region peptides, tagged with cell-permeable short peptides, greatly decrease Arf6 activation at growth cones in a time-dependent manner. Collectively, the signaling through CYTH2 and ACTN1 properly regulates neurite extension in N1E-115 cells, demonstrating the unexpected interaction of CYTH2 and ACTN1 in the regulation of cellular Arf6 activity involved in neurite extension. 相似文献
497.
Tomomura M Hasegawa H Suda N Sakagami H Tomomura A 《The Journal of biological chemistry》2012,287(22):17963-17974
Osteoclasts are essential for bone dynamics and calcium homeostasis. Recently, we reported that serum calcium-decreasing factor, caldecrin, which is a secretory-type serine protease isolated from the pancreas, inhibits osteoclast differentiation by suppression of NFATc1 activity regardless of its own protease activity (Hasegawa, H., Kido, S., Tomomura, M., Fujimoto, K., Ohi, M., Kiyomura, M., Kanegae, H., Inaba, A., Sakagami, H., and Tomomura, A. (2010) Serum calcium-decreasing factor, caldecrin, inhibits osteoclast differentiation by suppression of NFATc1 activity. J. Biol. Chem. 285, 25448-25457). Here, we investigated the effects of caldecrin on the function of mature osteoclasts by treatment with receptor activator of NF-κB ligand (RANKL). Caldecrin inhibited the RANKL-stimulated bone resorptive activity of mature osteoclasts. Furthermore, caldecrin inhibited RANKL-mediated sealing actin ring formation, which is associated with RANKL-evoked Ca(2+) entry through transient receptor potential vanilloid channel 4. The inhibitors of phospholipase Cγ, Syk, and c-Src suppressed RANKL-evoked Ca(2+) entry and actin ring formation of mature osteoclasts. Interestingly, caldecrin significantly inhibited RANKL-stimulated phosphorylation of c-Src, Syk, phospholipase Cγ1 and Cγ2, SLP-76, and Pyk2 but not that of ERK, JNK, or Akt. Caldecrin inhibited RANKL-stimulated c-Src kinase activity and c-Src·Syk association. These results suggest that caldecrin inhibits RANKL-stimulated calcium signaling activation and cytoskeletal organization by suppression of the c-Src·Syk pathway, which may in turn reduce the bone resorptive activity of mature osteoclasts. Thus, caldecrin is capable of acting as a negative regulator of osteoclastogenesis and osteoclast function of bone resorption. 相似文献
498.
499.
Kadotani A Fujimura M Nakamura T Ohyama S Harada N Maruki H Tamai Y Kanatani A Eiki J Nagata Y 《Archives of biochemistry and biophysics》2007,466(2):283-289
To investigate the effect of elevation of liver glycogen synthase (GYS2) activity on glucose and glycogen metabolism, we performed adenoviral overexpression of the mutant GYS2 with six serine-to-alanine substitutions in rat primary hepatocytes. Cell-free assays demonstrated that the serine-to-alanine substitutions caused constitutive activity and electrophoretic mobility shift. In rat primary hepatocytes, overexpression of the mutant GYS2 significantly reduced glucose production by 40% and dramatically induced glycogen synthesis via the indirect pathway rather than the direct pathway. Thus, we conclude that elevation of glycogen synthase activity has an inhibitory effect on glucose production in hepatocytes by shunting gluconeogenic precursors into glycogen. In addition, although intracellular compartmentation of glucose-6-phosphate (G6P) remains unclear in hepatocytes, our results imply that there are at least two G6P pools via gluconeogenesis and due to glucose phosphorylation, and that G6P via gluconeogenesis is preferentially used for glycogen synthesis in hepatocytes. 相似文献
500.
Hosoda C Hiroyama M Sanbe A Birumachi J Kitamura T Cotecchia S Simpson PC Tsujimoto G Tanoue A 《American journal of physiology. Heart and circulatory physiology》2007,293(1):H514-H519
Attenuation of early restenosis after percutaneous coronary intervention (PCI) is important for the successful treatment of coronary artery disease. Some clinical studies have shown that hypertension is a risk factor for early restenosis after PCI. These findings suggest that alpha(1)-adrenergic receptors (alpha(1)-ARs) may facilitate restenosis after PCI because of alpha(1)-AR's remarkable contribution to the onset of hypertension. In this study, we examined the neointimal formation after vascular injury in the femoral artery of alpha(1A)-knockout (alpha(1A)-KO), alpha(1B)-KO, alpha(1D)-KO, alpha(1A)-/alpha(1B)-AR double-KO (alpha(1AB)-KO), and wild-type mice to investigate the functional role of each alpha(1)-AR subtype in neointimal formation, which is known to promote restenosis. Neointimal formation 4 wk after wire injury was significantly (P < 0.05) smaller in alpha(1AB)-KO mice than in any other group of mice, while blood pressures were not altered in any of the groups of mice after wire injury compared with those before it. These results suggest that lack of both alpha(1A)- and alpha(1B)-ARs could be necessary to inhibit neointimal formation in the mouse femoral artery. 相似文献