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971.
Håkansson P Dahl L Chilkova O Domkin V Thelander L 《The Journal of biological chemistry》2006,281(3):1778-1783
Ribonucleotide reductase (RNR) is an essential enzyme that provides the cell with a balanced supply of deoxyribonucleoside triphosphates for DNA replication and repair. Mutations that affect the regulation of RNR in yeast and mammalian cells can lead to genetic abnormalities and cell death. We have expressed and purified the components of the RNR system in fission yeast, the large subunit Cdc22p, the small subunit Suc22p, and the replication inhibitor Spd1p. It was proposed (Liu, C., Powell, K. A., Mundt, K., Wu, L., Carr, A. M., and Caspari, T. (2003) Genes Dev. 17, 1130-1140) that Spd1 is an RNR inhibitor, acting by anchoring the Suc22p inside the nucleus during G1 phase. Using in vitro assays with highly purified proteins we have demonstrated that Spd1 indeed is a very efficient inhibitor of fission yeast RNR, but acting on Cdc22p. Furthermore, biosensor technique showed that Spd1p binds to the Cdc22p with a KD of 2.4 microM, whereas the affinity to Suc22p is negligible. Therefore, Spd1p inhibits fission yeast RNR activity by interacting with the Cdc22p. Similar to the situation in budding yeast, logarithmically growing fission yeast increases the dNTP pools 2-fold after 3 h of incubation in the UV mimetic 4-nitroquinoline-N-oxide. This increase is smaller than the increase observed in budding yeast but of the same order as the dNTP pool increase when synchronous Schizosaccharomyces pombe cdc10 cells are going from G1 to S-phase. 相似文献
972.
Haworth NL Feng LL Wouters MA 《Journal of bioinformatics and computational biology》2006,4(1):155-168
Redox-active disulfides are capable of being oxidized and reduced under physiological conditions. The enzymatic role of redox-active disulfides in thiol-disulfide reductases is well-known, but redox-active disulfides are also present in non-enzymatic protein structures where they may act as switches of protein function. Here, we examine disulfides linking adjacent beta-strands (cross-strand disulfides), which have been reported to be redox-active. Our previous work has established that these cross-strand disulfides have high torsional energies, a quantity likely to be related to the ease with which the disulfide is reduced. We examine the relationship between conformations of disulfides and their location in protein secondary structures. By identifying the overlap between cross-strand disulfides and various conformations, we wish to address whether the high torsional energy of a cross-strand disulfide is sufficient to confer redox activity or whether other factors, such as the presence of the cross-strand disulfide in a strained beta-sheet, are required. 相似文献
973.
小麦TaEDR1基因dsRNAi表达载体的构建及遗传转化 总被引:3,自引:0,他引:3
从抗白粉病小麦(TriticumaestivumL.)品系99/2439中分离到一个编码促分裂原蛋白激酶激酶激酶的新基因TaEDR1。为了研究TaEDR1基因在小麦抗白粉菌(Blumeriagraminis(DC.)E.O.Speerf.sp.triticiEm.Marchal)反应中的作用,以单子叶植物高效表达载体pAHC25为基础载体,选择TaEDR1基因编码胞外结构域的、保守性低的cDNA区域作为RNA干扰的目标区段,将这个区段连接成一个反向重复序列(3′→5′//5′←3′),插入到玉米泛素高效启动子Ubi1的下游,构建成双链RNA干扰表达载体pAH-R-TER。利用花粉管通道技术,以高产小麦新品种“周麦18”为受体进行了遗传转化。T0代植株经PPT抗性鉴定、PCR扩增检测,获得了7个转基因植株,为研究小麦TaEDR1基因的功能奠定了基础。 相似文献
974.
Aristoteli LP O'Rourke JL Danon S Larsson H Mellgard B Mitchell H Lee A 《Helicobacter》2006,11(5):460-468
BACKGROUND: Helicobacter pylori is a causative agent of gastric and duodenal ulcers and gastric cancer. Its urease enzyme allows survival in acid conditions and drives bacterial intracellular metabolism. We aimed to investigate the role of urease in determining the intragastric distribution of Helicobacter species in vivo. MATERIALS AND METHODS: The C57BL/6 mouse model of gastritis was used for infection with Helicobacter felis (CS1) or H. pylori (SS1). Urease-modulating compounds urea and/or fluorofamide (urease inhibitor) were administered to mice over 7 days. Concurrent gastric acid inhibition by omeprazole was also examined. Bacterial distribution in the antrum, body, antrum/body, and body/cardia transitional zones was graded "blindly" by histologic evaluation. Bacterial colony counts on corresponding tissue were also conducted. RESULTS: Urease inhibition by fluorofamide decreased H. pylori survival in most gastric regions (p < .05); however, there were no marked changes to H. felis colonization after this treatment. There was a consistent trend for decreased antral colonization, and an increase in antrum/body transitional zone and body colonization with excess 5% or 6% (w/v) urea treatment. Significant reductions of both Helicobacter species were observed with the co-treatment of urea and fluorofamide (p < .05). Collateral treatment with omeprazole did not alter H. pylori colonization patterns caused by urea/fluorofamide. CONCLUSIONS: Urease perturbations affect colonization patterns of Helicobacter species. Combined urea and fluorofamide treatment reduced the density of both Helicobacter species in our infection model. 相似文献
975.
Silva LC Oliveira LS Mendes GD Garcia G Pereira Ados S De Nucci G 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2006,832(2):302-306
Isosorbide 5-mononitrate (5-ISMN) is an organic nitrate widely used for its vasodilating properties in the treatment of angina pectoris. In the present study, an efficient, sensitive, robust method was developed for the determination and quantification of isosorbide 5-mononitrate, in human plasma, by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS), using photospray ionization. Isosorbide 5-mononitrate was extracted from 0.5 mL human plasma by liquid-liquid extraction (LLE). The method had a chromatographic run of 2.0 min using a C(8) analytical column (100 mm x 2.1 mm i.d.) and the linear calibration curve over the range was linear from 20 to 2000 ng mL(-1) (r(2)>0.995). The between-run precision, based on the relative standard deviation replicate quality controls, was 7.9% (60 ng mL(-1)), 5.2% (300 ng mL(-1)) and 7.0% (1800 ng mL(-1)). The between-run accuracy was 94.9%, 94.1% and 88.8% for the above-mentioned concentrations, respectively. The method herein described was employed in a bioequivalence study of two tablet formulations of isosorbide 5-mononitrate 40 mg. 相似文献
976.
Sehon C McClure K Hack M Morton M Gomez L Li L Barrett TD Shankley N Breitenbucher JG 《Bioorganic & medicinal chemistry letters》2006,16(1):77-80
High-throughput screening revealed compound 1 as a potent antagonist of the CCK(1) receptor. Here, we disclose the synthesis of combinatorial libraries by solid-phase synthesis on Kenner 'safety catch' resin. Additive QSAR models were used to determine a lack of consistent additive SAR within the matrix. 相似文献
977.
978.
A linear water-insoluble (1-->3)-beta-D-glucan, coded as GL-IV-I, was isolated from the fruit body of Ganoderma lucidum by extracting with NaOH solution. Its derivatives were prepared by using sulfation, carboxymethylation, hydroxyethylation, hydroxypropylation, and methylation, respectively, and these were labeled as S-GL, CM-GL, HE-GL, HP-GL and M-GL. Five derivatives exhibited good water solubility. Their structures and chain conformations were investigated with infrared spectroscopy, elemental analysis (EA), one- and two-dimensional NMR spectroscopy, laser light scattering (LLS), and size-exclusion chromatography combined with LLS (SEC-LLS). The reactivity of the hydroxyl group of GL-IV-I was ordered as C-6>C-4>C-2 for the five derivatives. The degree of substitution (DS) of the derivatives was calculated from EA and NMR spectroscopy to be from 0.32 to 1.18. The weight-average molecular mass (M(w)) of GL-IV-I, S-GL, CM-GL, HE-GL, HP-GL, and M-GL was 13.3 x 10(4), 10.1 x 10(4), 6.3 x 10(4), 7.2 x 10(4), 5.1 x 10(4), and 14.1 x 10(4), respectively. The conformation analysis studies revealed that GL-IV-I exists as a compact coil in dimethyl sulfoxide, whereas the five derivatives are slightly expanded flexible chains in 0.9% aqueous NaCl solution. 相似文献
979.
Physicochemical properties and antitumor activities for sulfated derivatives of lentinan 总被引:1,自引:0,他引:1
Five fractions of lentinan, a β-(1→3)-d-glucan bearing β-(1→6)-d-glucopyranosyl branches, were treated with chlorosulfonic acid for 90 min at 60 °C in pyridine medium to synthesize water-soluble sulfated derivatives having the substitution degree of 1.44–1.76. The 13C NMR spectra of the sulfated β-glucans indicated that the C-6 position was preferentially substituted by the sulfate groups. The values of the weight-average molecular weight (Mw), radius of gyration (), and intrinsic viscosity ([η]) of the sulfated lentinan fractions were determined by size-exclusion chromatography with multi-angle laser light scattering (SEC–MALLS) and viscometry in 0.15 M aq NaCl at 25 °C, respectively. The dependence of [η] on Mw for the sulfated lentinan was found to be [η] = 8.93 × 10−3 (mL/g) in 0.15 M aq NaCl (for Mw ranging from 14.6 × 104 to 50.4 × 104). On the basis of the Yamakawa–Fujii–Yoshizaki (YFY) theory, the conformational parameters of the sulfated lentinan were calculated as 950 nm−1 for the molar mass per unit contour length (ML), 4.8 nm for the persistence length (q), and 13.9 for the characteristic ratio (C∞), indicating relatively extended single flexible chains in solution. The sulfated glucan fractions exhibited in vitro antiproliferative activities against sarcoma 180 (S-180) cells, and their inhibition ratios were lower than that of the triple-helix lentinan, but higher than that for the one with single random-coil lentinan chains. 相似文献
980.
Chun Shi Lina Zhao Bilian Zhu Qianxi Li David T. Yew Zhibin Yao Jie Xu 《Chemico-biological interactions》2009,180(3):389-397
Standardized extract from the leaves of the Ginkgo biloba tree, labeled EGb761, is one of the most popular herbal supplements, taken for its multivalent properties. In this study, dosage effects of EGb761 on hydrogen peroxide (H2O2)-induced apoptosis of human neuroblastoma SH-SY5Y cells were investigated. It was found that H2O2-induced apoptotic cell death in SH-SY5Y cells, which was revealed in DNA fragmentation, mitochondrial membrane potential depolarization, and activation of Akt, c-Jun N-terminal kinases (JNK) and caspase 3. Low doses of EGb761 (50–100 μg/ml) inhibited H2O2-induced cell apoptosis via inactivation of Akt, JNK and caspase 3 while high doses of EGb761 (250–500 μg/ml) enhanced H2O2 toxicities via inactivation of Akt and enhancement of activation of JNK and caspase 3. Additional experiments revealed that H2O2 decreased intracellular GSH content, which was also inhibited by low concentrations of EGb761 but enhanced after high concentrations of EGb761 treatment. This further suggests to us that dosage effects of EGb761 on apoptotic signaling proteins may be correlated with regulation of cell redox state. Therefore, treatment dosage may be one of the vital factors that determine the specific action of EGb761 on oxidative stress-induced cell apoptosis. To understand the mechanisms of dosage effects of EGb761 may have important clinical implications. 相似文献