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211.
Furukawa R Yamada Y Takenaga M Igarashi R Harashima H 《Biochemical and biophysical research communications》2011,404(3):796-801
The anti-oxidant enzyme superoxide dismutase (SOD) has the potential for use as a therapeutic agent in the treatment of various diseases caused by reactive oxygen species. However, achieving this would be difficult without a suitable delivery system for SOD. We previously reported that PC-SOD, in which four molecules of a phosphatidylcholine (PC) derivative were covalently bound to each dimer of recombinant human CuZnSOD, was a high affinity for the cell membrane [14]. Here, we show that an octaarginine (R8) modified liposome equipped with PC-SOD (R8-LP (PC-SOD)) enhances its anti-oxidant effect. High-density R8-modified liposomes can stimulate macropinocytosis and are taken up efficiently by cells as demonstrated in a previous study [21]. Flow cytometry analyses showed that R8-LP (PC-SOD) was taken up by cells more efficiently than PC-SOD. Moreover, R8-LP (PC-SOD) liposomes were found to scavenge superoxide anions (O2−) very efficiently. These results suggest that the efficient cytosolic delivery of PC-SOD by R8-modified liposomes would enhance the anti-oxidant effects of PC-SOD. 相似文献
212.
8-Oxo-7,8-dihydroguanine (8-hydroxyguanine) is oxidized more easily than normal nucleobases, which can produce spiroiminodihydantoin (Sp) and guanidinohydantoin (Gh). These secondary oxidation products of 8-oxo-7,8-dihydroguanine are highly mutagenic when formed within DNA. To evaluate the mutagenicity of the corresponding oxidation products of 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphate (8-hydroxy-2'- deoxyguanosine 5'-triphosphate) in the nucleotide pool, Escherichia coli cells deficient in the mutT gene were treated with H(2)O(2), and the induced mutations were analyzed. Moreover, the 2'-deoxyriboside 5'-triphosphate derivatives of Sp and Gh were also introduced into competent E. coli cells. The H(2)O(2) treatment of mutT E. coli cells resulted in increase of G:C → T:A and A:T → T:A mutations. However, the incorporation of exogenous Sp and Gh 2'-deoxyribonucleotides did not significantly increase the mutation frequency. These results suggested that the oxidation product(s) of 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphate induces G:C → T:A and A:T → T:A mutations, and that the 2'-deoxyriboside 5'-triphosphate derivatives of Sp and Gh exhibit quite weak mutagenicity, in contrast to the bases in DNA. 相似文献
213.
Zheng Zheng Teruo Nonomura Michela Appiano Stefano Pavan Yoshinori Matsuda Hideyoshi Toyoda Anne-Marie A. Wolters Richard G. F. Visser Yuling Bai 《PloS one》2013,8(7)
Powdery mildew disease caused by Leveillula taurica is a serious fungal threat to greenhouse tomato and pepper production. In contrast to most powdery mildew species which are epiphytic, L. taurica is an endophytic fungus colonizing the mesophyll tissues of the leaf. In barley, Arabidopsis, tomato and pea, the correct functioning of specific homologues of the plant Mlo gene family has been found to be required for pathogenesis of epiphytic powdery mildew fungi. The aim of this study was to investigate the involvement of the Mlo genes in susceptibility to the endophytic fungus L. taurica. In tomato (Solanum lycopersicum), a loss-of-function mutation in the SlMlo1 gene results in resistance to powdery mildew disease caused by Oidium neolycopersici. When the tomato Slmlo1 mutant was inoculated with L. taurica in this study, it proved to be less susceptible compared to the control, S. lycopersicum cv. Moneymaker. Further, overexpression of SlMlo1 in the tomato Slmlo1 mutant enhanced susceptibility to L. taurica. In pepper, the CaMlo2 gene was isolated by applying a homology-based cloning approach. Compared to the previously identified CaMlo1 gene, the CaMlo2 gene is more similar to SlMlo1 as shown by phylogenetic analysis, and the expression of CaMlo2 is up-regulated at an earlier time point upon L. taurica infection. However, results of virus-induced gene silencing suggest that both CaMlo1 and CaMlo2 may be involved in the susceptibility of pepper to L. taurica. The fact that overexpression of CaMlo2 restored the susceptibility of the tomato Slmlo1 mutant to O. neolycopersici and increased its susceptibility to L. taurica confirmed the role of CaMlo2 acting as a susceptibility factor to different powdery mildews, though the role of CaMlo1 as a co-factor for susceptibility cannot be excluded. 相似文献
214.
215.
Takeuchi M Yoshioka H Seo Y Tanabe S Tamaki H Kamagata Y Takahashi HA Igari S Mayumi D Sakata S 《Environmental microbiology》2011,13(12):3206-3218
Anaerobic methane-oxidizing archaea (ANME) are known to play an important role in methane flux, especially in marine sediments. The 16S rRNA genes of ANME have been detected in terrestrial freshwater subsurfaces. However, it is unclear whether ANME are actively involved in methane oxidation in these environments. To address this issue, Holocene sediments in the subsurface of the Kanto Plain in Japan were collected for biogeochemical and molecular analysis. The potential activity of the anaerobic oxidation of methane (AOM) (0.38-3.54 nmol cm?3 day?1) was detected in sediment slurry incubation experiments with a (13) CH(4) tracer. Higher AOM activity was observed in low-salinity treatment compared with high-salinity condition (20‰), which supports the adaptation of ANME in freshwater habitats. The 16S rRNA sequence analysis clearly revealed the presence of a distinct subgroup of ANME-1, designated ANME-1a-FW. Phylogenetic analysis of the mcrA genes also implied the presence of the distinct subgroup in ANME-1. ANME-1a-FW was found to be the most dominant active group in the archaeal communities on the basis of 16S rRNA analysis (75.0-93.8% of total archaeal 16S rRNA clones). Sulfate-reducing bacteria previously known as the syntrophic bacterial partners of ANME-1 was not detected. Our results showed that ANME-1a-FW is adapted to freshwater habitats and is responsible for AOM in terrestrial freshwater subsurface environments. 相似文献
216.
Hikaru Hemmi Takashi Kumazaki Shuichi Kojima Takuya Yoshida Tadayasu Ohkubo Hideyoshi Yokosawa Yuji Kobayashi 《Journal of peptide science》2011,17(8):595-600
P14C/N39C is the disulfide variant of the ovomucoid third domain from silver pheasant (OMSVP3) introducing an engineered Cys14? Cys39 bond near the reactive site on the basis of the sequence homology between OMSVP3 and ascidian trypsin inhibitor. This variant exhibits a narrower inhibitory specificity. We have examined the effects of introducing a Cys14? Cys39 bond into the flexible N‐terminal loop of OMSVP3 on the thermodynamics of the reactive site peptide bond hydrolysis, as well as the thermal stability of reactive site intact inhibitors. P14C/N39C can be selectively cleaved by Streptomyces griseus protease B at the reactive site of OMSVP3 to form a reactive site modified inhibitor. The conversion rate of intact to modified P14C/N39C is much faster than that for wild type under any pH condition. The pH‐independent hydrolysis constant (Khyd°) is estimated to be approximately 5.5 for P14C/N39C, which is higher than the value of 1.6 for natural OMSVP3. The reactive site modified form of P14C/N39C is thermodynamically more stable than the intact one. Thermal denaturation experiments using intact inhibitors show that the temperature at the midpoint of unfolding at pH 2.0 is 59 °C for P14C/N39C and 58 °C for wild type. There have been no examples, except P14C/N39C, where introducing an engineered disulfide causes a significant increase in Khyd°, but has no effect on the thermal stability. The site‐specific disulfide introduction into the flexible N‐terminal loop of natural Kazal‐type inhibitors would be useful to further characterize the thermodynamics of the reactive site peptide bond hydrolysis. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
217.
Abe N Abe H Nagai C Harada M Hatakeyama H Harashima H Ohshiro T Nishihara M Furukawa K Maeda M Tsuneda S Ito Y 《Bioconjugate chemistry》2011,22(10):2082-2092
RNA interference (RNAi) is one of the most promising new approaches for disease therapy. The design of a dumbbell-shaped nanocircular RNA allows it to act as a short interfering RNA (siRNA) precursor. To optimize the design, we studied the relationship between the nanostructure and RNAi activity by synthesizing various RNA dumbbells. An RNA dumbbell with a 23-bp stem and 9-nt loops was the most potent. Sequence analysis by mass spectrometry showed that Dicer could edit RNA dumbbells to siRNA species. The reaction offered the slow release of siRNA species, which conferred prolonged RNAi activity. Introduction of DNA into the loop position significantly stabilized the dumbbell in biological fluid without any loss of RNAi activity. In-depth pharmacological evaluation was performed by introducing dumbbells into HeLa cells that stably express the target luciferase gene. The dumbbells provided a rapid silencing effect and retained this effect for a longer time even at a lower concentration than that at which standard siRNA completely lost RNAi activity. We conclude that an RNA dumbbell with DNA loops is the most promising design for in vivo applications for RNA medicine. 相似文献
218.
The intranuclear disposition of a plasmid is extremely important for transgene expression. The effects of a left-handedly curved sequence with high histone affinity on plasmid expression were examined in vivo. A naked luciferase-plasmid was delivered into mouse liver by a hydrodynamics-based injection, and the luciferase activities were quantitated at various time points. The location of the left-handedly curved sequence determined the transgene expression, without affecting the amount of intranuclear exogenous DNA. The plasmid containing the curved sequence at the location that results in the exposure of the TATA box out of the nucleosome core showed the highest expression. These results suggest that sequences with high histone affinity could control transgene expression from plasmids in vivo. 相似文献
219.
Microbial diversity and methanogenic potential in a high temperature natural gas field in Japan 总被引:1,自引:0,他引:1
Mochimaru H Yoshioka H Tamaki H Nakamura K Kaneko N Sakata S Imachi H Sekiguchi Y Uchiyama H Kamagata Y 《Extremophiles : life under extreme conditions》2007,11(3):453-461
Microbial diversity and methanogenic potential in formation water samples from a dissolved-in-water type gas field were investigated
by using 16S rRNA gene libraries and culture-based methods. Two formation water samples (of 46 and 53°C in temperature) were
obtained from a depth of 700 to 800 m. Coenzyme F420-autofluorescence indicated that 103–104 cells per ml of active methanogens were present, accounting for at least 10% of the total cell count. The 16S rRNA gene sequence
analysis indicated that the diversity of Archaea and Bacteria of the two samples was quite limited; i.e., the archaeal libraries were dominated by the sequences related to Methanobacterium
formicicum and Methanothermobacter thermautotrophicus, and the bacterial libraries were dominated by the sequences related to Hydrogenophilus and Deferribacter. Of the methanogenic substrates tested using the formation water-based medium, only H2–CO2 gave rise to methane formation. Those dominant archaeal and bacterial genera have the potential to use hydrogen for growth
at the in situ temperatures, suggesting that the formation water of the Pliocene strata in the gas field has been provided
with hydrogen, probably from underneath the strata, and thus on-going active methanogenesis has been occurring to date.
An erratum to this article can be found at 相似文献
220.
Recent progress in genetics and molecular biology has provided useful information regarding the molecular mechanisms associated with the mitochondrial diseases. Genetic approaches were initiated in the late 1980s to clarify the gene responsible for various mitochondrial diseases, and information concerning genetic mutations is currently used in the diagnosis of mitochondrial diseases. Moreover, it was also revealed that mitochondria play a central role in apoptosis, or programmed cell death, which is closely related to the loss of physiological functions of tissues. Therefore, drug therapies targeted to the mitochondria would be highly desirable. In spite of the huge amount of mechanism-based studies of mitochondrial diseases, effective therapies have not yet been established mainly because of the lack of an adequate delivery system. To date, numerous investigators have attempted to establish a mitochondrial drug delivery system. However, many problems remain to be overcome before a clinical application can be achieved. To fulfill a drug delivery targeted to mitochondria, we first need to establish a method to encapsulate various drugs, proteins, peptides, and genes into a drug carrier depending on their physical characteristics. Second, we need to target it to a specific cell. Finally, multi-processes of intracellular trafficking should be sophisticatedly regulated so as to release a drug carrier from the endosome to the cytosol, and thereafter to deliver to the mitochondria. In this review, we describe the current state of the development of mitochondrial drug delivery systems, and discuss the advantage and disadvantage of each system. Our current efforts to develop an efficient method for the packaging of macromolecules and regulating intracellular trafficking are also summarized. Furthermore, novel concept of "Regulation of intramitochondrial trafficking" is proposed herein as a future challenge to the development of a mitochondrial drug delivery system. 相似文献