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971.
Guoyong Hu Jiaqin Shen Li Cheng Di Xiang Zhonghui Zhang Miao He Huili Lu Shunying Zhu Mingyuan Wu Yan Yu Xingpeng Wang Wei Han 《Protein expression and purification》2010,69(2):186-190
Regenerating gene (Reg) IV is a newly discovered member of the regenerating gene family belonging to the calcium (C-type) dependent lectin superfamily. Reg IV is highly expressed in the gastrointestinal tract and markedly up-regulated in colon adenocarcinoma, pancreatic cancer, gastric adenocarcinoma, and inflammatory bowel disease. However, the physiological and pathological functions of Reg IV are largely unknown, partly due to the limited access of the bioactive protein. We report here the first expression and purification of Reg IV proteins using a prokaryotic system. Human Reg IV was expressed in Escherichia coli as an insoluble protein which was identified in the fraction of inclusion body after ultrasonication of the bacteria. After the protein aggregate was solubilized by guanidine–HCl, it was refolded by sucrose and arginine-assisted procedures and purified using cation-exchange chromatography. The protein identity and purity of the final preparation were confirmed by analysis of the protein mass and immune specificity in SDS–PAGE, Western blotting, and HPLC assay. The biological activity of the protein was determined by the HCT116 and HT29 cell proliferation assays. The highly purified bioactive human Reg IV should aid in further characterization of its physiological and pathological functions. 相似文献
972.
Dai H Shin OH Machius M Tomchick DR Südhof TC Rizo J 《Nature structural & molecular biology》2004,11(9):844-849
The neuronal protein synaptotagmin 1 functions as a Ca(2+) sensor in exocytosis via two Ca(2+)-binding C(2) domains. The very similar synaptotagmin 4, which includes all the predicted Ca(2+)-binding residues in the C(2)B domain but not in the C(2)A domain, is also thought to function as a neuronal Ca(2+) sensor. Here we show that, unexpectedly, both C(2) domains of fly synaptotagmin 4 exhibit Ca(2+)-dependent phospholipid binding, whereas neither C(2) domain of rat synaptotagmin 4 binds Ca(2+) or phospholipids efficiently. Crystallography reveals that changes in the orientations of critical Ca(2+) ligands, and perhaps their flexibility, render the rat synaptotagmin 4 C(2)B domain unable to form full Ca(2+)-binding sites. These results indicate that synaptotagmin 4 is a Ca(2+) sensor in the fly but not in the rat, that the Ca(2+)-binding properties of C(2) domains cannot be reliably predicted from sequence analyses, and that proteins clearly identified as orthologs may nevertheless have markedly different functional properties. 相似文献
973.
Duk-Young Jung Takashi Yamada Toshie Tsuchiya Su-Chak Ryu Dong-Wook Han 《Biotechnology and Bioprocess Engineering》2010,15(2):341-348
Many studies have been performed to accelerate osteoinduction and osteoconduction into porous ceramic scaffolds by seeding
them with cells. In this study, we compared available cell-seeding methods on a porous β-tricalcium phosphate (β-TCP) scaffold
and evaluated the effects of cell-seeding on the mechanical properties of the porous β-TCP scaffold. Three types of porous
bioceramic scaffolds were used: dry scaffold, scaffold wetted with media, and scaffold cultivated with normal human osteoblasts
(NHOs). Cell-seeding into the porous β-TCP scaffolds was performed by conventional, centrifuge, high-density, and vacuum methods.
After confirming cell proliferation with MTT assay and cell staining, a compressive test was performed after 2 and 4 weeks
of cell culture. The vacuum method based on the high-density cell culture inserted effectively NHOs into the β-TCP scaffolds.
The compressive elastic modulus of wetted β-TCP scaffolds decreased significantly (p < 0.05) about 20∼30% after 2 and 4 weeks of incubation in comparison with that of the dry scaffold. However, the compressive
strength of the scaffolds cultivated with NHOs for 3 weeks was significantly (p < 0.05) higher than that of scaffolds without NHOs. The vacuum with the high-density of cell-seeding seems to be a suitable
method for seeding cells into complex porous ceramic scaffolds. Cell proliferation and uniform distribution in the scaffolds
can change the initial mechanical properties of porous ceramic scaffolds. 相似文献
974.
Fusicoccin (FC) treatment prevents dark‐induced stomatal closure, the mechanism of which is still obscure. By using pharmacological approaches and laser‐scanning confocal microscopy, the relationship between FC inhibition of dark‐induced stomatal closure and the hydrogen peroxide (H2O2) levels in guard cells in broad bean was studied. Like ascorbic acid (ASA), a scavenger of H2O2 and diphenylene iodonium (DPI), an inhibitor of H2O2‐generating enzyme NADPH oxidase, FC was found to inhibit stomatal closure and reduce H2O2 levels in guard cells in darkness, indicating that FC‐caused inhibition of dark‐induced stomatal closure is related to the reduction of H2O2 levels in guard cells. Furthermore, like ASA, FC not only suppressed H2O2‐induced stomatal closure and H2O2 levels in guard cells treated with H2O2 in light, but also reopened the stomata which had been closed by darkness and reduced the level of H2O2 that had been generated by darkness, showing that FC causes H2O2 removal in guard cells. The butyric acid treatment simulated the effects of FC on the stomata treated with H2O2 and had been closed by dark, and on H2O2 levels in guard cells of stomata treated with H2O2 and had been closed by dark, and both FC and butyric acid reduced cytosol pH in guard cells of stomata treated with H2O2 and had been closed by dark, which demonstrates that cytosolic acidification mediates FC‐induced H2O2 removal. Taken together, our results provide evidence that FC causes cytosolic acidification, consequently induces H2O2 removal, and finally prevents dark‐induced stomatal closure. 相似文献
975.
Han J Hou J Liu H Cai S Feng B Zhou J Xiang H 《Applied and environmental microbiology》2010,76(23):7811-7819
Polyhydroxyalkanoates (PHAs) are accumulated as intracellular carbon and energy storage polymers by various bacteria and a few haloarchaea. In this study, 28 strains belonging to 15 genera in the family Halobacteriaceae were investigated with respect to their ability to synthesize PHAs and the types of their PHA synthases. Fermentation results showed that 18 strains from 12 genera could synthesize polyhydroxybutyrate (PHB) or poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV). For most of these haloarchaea, selected regions of the phaE and phaC genes encoding PHA synthases (type III) were cloned via PCR with consensus-degenerate hybrid oligonucleotide primers (CODEHOPs) and were sequenced. The PHA synthases were also examined by Western blotting using haloarchaeal Haloarcula marismortui PhaC (PhaC(Hm)) antisera. Phylogenetic analysis showed that the type III PHA synthases from species of the Halobacteriaceae and the Bacteria domain clustered separately. Comparison of their amino acid sequences revealed that haloarchaeal PHA synthases differed greatly in both molecular weight and certain conserved motifs. The longer C terminus of haloarchaeal PhaC was found to be indispensable for its enzymatic activity, and two additional amino acid residues (C143 and C190) of PhaC(Hm) were proved to be important for its in vivo function. Thus, we conclude that a novel subtype (IIIA) of type III PHA synthase with unique features that distinguish it from the bacterial subtype (IIIB) is widely distributed in haloarchaea and appears to be involved in PHA biosynthesis. 相似文献
976.
Göker M Held B Lapidus A Nolan M Spring S Yasawong M Lucas S Glavina Del Rio T Tice H Cheng JF Goodwin L Tapia R Pitluck S Liolios K Ivanova N Mavromatis K Mikhailova N Pati A Chen A Palaniappan K Brambilla E Land M Hauser L Chang YJ Jeffries CD Brettin T Detter JC Han C Rohde M Sikorski J Woyke T Bristow J Eisen JA Markowitz V Hugenholtz P Kyrpides NC Klenk HP 《Standards in genomic sciences》2010,3(1):66-75
Ignisphaera aggregans Niederberger et al. 2006 is the type and sole species of genus Ignisphaera. This archaeal species is characterized by a coccoid-shape and is strictly anaerobic, moderately acidophilic, heterotrophic hyperthermophilic and fermentative. The type strain AQ1.S1(T) was isolated from a near neutral, boiling spring in Kuirau Park, Rotorua, New Zealand. This is the first completed genome sequence of the genus Ignisphaera and the fifth genome (fourth type strain) sequence in the family Desulfurococcaceae. The 1,875,953 bp long genome with its 2,009 protein-coding and 52 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project. 相似文献
977.
978.
Heparan Sulfate D‐Glucosaminyl 3‐O‐Sulfotransferase‐3B1 (HS3ST3B1) Promotes Angiogenesis and Proliferation by Induction of VEGF in Acute Myeloid Leukemia Cells 下载免费PDF全文
Lei Zhang Kai Song Ling Zhou Zhishen Xie Ping Zhou Yiming Zhao Yue Han Xiaojun Xu Ping Li 《Journal of cellular biochemistry》2015,116(6):1101-1112
979.
980.
Weirong Fang Rui Zhang Lan Sha Peng Lv Erxin Shang Dan Han Jie Wei Xiaohan Geng Qichuan Yang Yunman Li 《Journal of neurochemistry》2014,128(5):662-671
The blood–brain barrier (BBB) greatly limits the efficacy of many neuroprotective drugs' delivery to the brain, so improving drug penetration through the BBB has been an important focus of research. Here we report that platelet activating factor (PAF) transiently opened BBB and facilitated neuroprotectant edaravone penetration into the brain. Intravenous infusion with PAF induced a transient BBB opening in rats, reflected by increased Evans blue leakage and mild edema formation, which ceased within 6 h. Furthermore, rat regional cerebral blood flow (rCBF) declined acutely during PAF infusion, but recovered slowly. More importantly, this transient BBB opening significantly increased the penetration of edaravone into the brain, evidenced by increased edaravone concentrations in tissue interstitial fluid collected by microdialysis and analyzed by Ultra‐performance liquid chromatograph combined with a hybrid quadrupole time‐of‐flight mass spectrometer (UPLC‐MS/MS). Similarly, incubation of rat brain microvessel endothelial cells monolayer with 1 μM PAF for 1 h significantly increased monolayer permeability to 125I‐albumin, which recovered 1 h after PAF elimination. However, PAF incubation with rat brain microvessel endothelial cells for 1 h did not cause detectable cytotoxicity, and did not regulate intercellular adhesion molecule‐1, matrix‐metalloproteinase‐9 and P‐glycoprotein expression. In conclusion, PAF could induce transient and reversible BBB opening through abrupt rCBF decline, which significantly improved edaravone penetration into the brain.