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991.
Yun Hee Choi Hye Jeong Choi Dooil Kim Ki-Nam Uhm Hyung-Kwoun Kim 《Applied microbiology and biotechnology》2010,87(1):185-193
3-Chloro-1-phenyl-1-propanol is used as a chiral intermediate in the synthesis of antidepressant drugs. Various microbial
reductases were expressed in Escherichia coli, and their activities toward 3-chloro-1-phenyl-1-propanone were evaluated. The yeast reductase YOL151W (GenBank locus tag)
exhibited the highest level of activity and exclusively generated the (S)-alcohol. Recombinant YOL151W was purified by Ni-nitrilotriacetic acid (Ni-NTA) and desalting column chromatography. It displayed
an optimal temperature and pH of 40°C and 7.5–8.0, respectively. The glucose dehydrogenase coupling reaction was introduced
as an NADPH regeneration system. NaOH solution was occasionally added to maintain the reaction solution pH within the range
of 7.0–7.5. By using this reaction system, the substrate (30 mM) could be completely converted to the (S)-alcohol product with an enantiomeric excess value of 100%. A homology model of YOL151W was constructed based on the structure
of Sporobolomyces salmonicolor carbonyl reductase (Protein Data Bank ID: 1Y1P). A docking model of YOL151W with NADPH and 3-chloro-1-phenyl-1-propanone
was then constructed, which showed that the cofactor and substrate bound tightly to the active site of the enzyme in the lowest
free energy state and explained how the (S)-alcohol was produced exclusively in the reduction process. 相似文献
992.
Man Deok Kim Minkyung Song Minho Jo Seung Gu Shin Jee Hyeong Khim Seokhwan Hwang 《Applied microbiology and biotechnology》2010,85(5):1611-1618
This paper reports the effects of changing pH (5–7) and temperature (T, 40–60 °C) on the efficiencies of bacterial hydrolysis of suspended organic matter (SOM) in wastewater from food waste recycling
(FWR) and the changes in the bacterial community responsible for this hydrolysis. Maximum hydrolysis efficiency (i.e., 50.5%
reduction of volatile suspended solids) was predicted to occur at pH 5.7 and T = 44.5 °C. Changes in short-chain volatile organic acid profiles and in acidogenic bacterial communities were investigated
under these conditions. Propionic and butyric acids concentrations increased rapidly during the first 2 days of incubation.
Several band sequences consistent with Clostridium spp. were detected using denaturing gel gradient electrophoresis. Clostridium thermopalmarium and Clostridium novyi seemed to contribute to butyric acid production during the first 1.5 days of acidification of FWR wastewater, and C. thermopalmarium was a major butyric acid producer afterward. C. novyi was an important propionic acid producer. These two species appear to be important contributors to hydrolysis of SOM in the
wastewater. Other acidogenic anaerobes, Aeromonas sharmana, Bacillus coagulans, and Pseudomonas plecoglossicida, were also indentified. 相似文献
993.
Choong Hyun Lee Jung Hoon Choi Ki-Yeon Yoo Ok Kyu Park In Koo Hwang Sang Guan You Boo-Yong Lee Il-Jun Kang Moo-Ho Won 《Cellular and molecular neurobiology》2010,30(2):255-263
Melatonin exerts many physiological functions via its G protein-coupled receptors. In the present study, we investigated age-related
changes in MT2 melatonin receptor immunoreactivity and its levels in the gerbil hippocampus during normal aging. In the postnatal
month 1 (PM 1) group, MT2 immunoreaction was well observed in neurons in all subregions of the gerbil hippocampus. In the
PM 3 and 6 groups, MT2 immunoreactivity in neurons was decreased compared to that in the PM 1 group. Thereafter, MT2 immunoreactivity
in neurons was increased. In the PM 18 and 24 groups, MT2 immunoreactivity in neurons was strong in all subregions of the
gerbil hippocampus. In addition, the number of MT2 immunoreactive cells was lowest at PM 3 and highest at PM 24. From western
blot analysis, age-dependent change pattern in MT2 level in the gerbil hippocampus was similar to the immunohistochemical
result. These results indicate that MT2 immunoreactivity and levels are altered in the gerbil hippocampus during normal aging;
lowest at young adult stage and highest at aged stage. 相似文献
994.
Eun-A Kim Hoh-Gyu Hahn Key-Sun Kim Tae Ue Kim Soo Young Choi Sung-Woo Cho 《Cellular and molecular neurobiology》2010,30(5):807-815
We have screened new drugs with a view to developing effective drugs against glutamate-induced excitotoxicity. In the present
work, we show effects of a new drug, 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride against glutamate-induced excitotoxicity
in primary rat glial cultures. Pretreatment of glial cells with 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride for
2 h significantly protected glial cells against glutamate-induced excitotoxicity in a time- and dose-dependent manner with
an optimum concentration of 100 μM. The drug significantly reduced production of proinflammatory cytokines, tumor necrosis
factor-α, and interlukin-1β in glutamate-induced excitotoxicity. The drug also prevented glutamate-induced intracellular Ca2+ influx and reduced the subsequent overproduction of nitric oxide and reactive oxygen species. Furthermore, the drug preserved
the mitochondrial potential and inhibited the overproduction of cytochrome c. In addition, the drug effectively attenuated the protein level changes of β-catenin and glycogen synthase kinase-3β. These
results suggest that 2-cyclopropylimino-3-methyl-1,3-thiazoline hydrochloride effectively protected primary cultures of rat
glial cells against glutamate-induced excitotoxicity. 相似文献
995.
Dae Won Chung Ki-Yeon Yoo In Koo Hwang Dae Won Kim Jin Young Chung Choong Hyun Lee Jung Hoon Choi Soo Young Choi Hwa Young Youn In Se Lee Moo-Ho Won 《Cellular and molecular neurobiology》2010,30(4):531-541
In this study, we observed the effects of lipopolysaccharide (LPS) on neurodegeneration and immune response in the hippocampus. LPS is a gram-negative bacterial cell surface proteoglycan and known as a bacterial endotoxin. For this, we investigated the optimal concentration of LPS influencing the ICR mouse hippocampus to measure the LPS receptor, e.g., toll-like receptor 4 (TLR4), expression in mouse hippocampal homogenates. TLR4 expression was significantly and prominently increased in the hippocampal homogenates of the LPS (1 mg/kg)-treated group. Next, we examined pro-inflammatory response in the hippocampus using cyclooxygenase-2 (COX-2, a marker for inflammatory response) immunohistochemistry after LPS treatment. COX-2 immunoreactivity was significantly increased in the endothelium of blood vessels in the hippocampus 6 h after LPS treatment, judging from double immunofluorescence study with platelet-derived endothelial cell adhesion molecule-1 (PECAM-1, a marker for endothelial cells): it decreased 12 h and disappeared 24 h after LPS treatment. In addition, the ionized calcium-binding adapter molecule 1 (Iba-1)-immunoreactive (+) microglia were morphologically activated in the mouse hippocampus after LPS treatment. At 24 h after LPS treatment, Iba-1+ microglia of activated forms were abundant in the hippocampus. However, NeuN (a neuron-specific soluble nuclear antigen)+ neurons were not significantly changed in the hippocampus after LPS treatment. Fluoro-jade B (a marker for neuronal degeneration)+ cells were not detected in the hippocampus at any time after LPS treatment. In addition, there were no significant differences in permeability of blood–brain barriers at any time points after LPS treatment. In brief, our results indicate that intraperitoneal administration of 1 mg/kg LPS effectively induces LPS receptor (TLR4) expression in the hippocampus, and the treatment increases corticosterone levels, inflammation in the blood vessels, and microglial activation in the hippocampus without any neuronal damage. 相似文献
996.
The inositol 1,4,5-trisphosphate (IP3)-mediated intracellular Ca2+ releases in secretory cells play vital roles in controlling not only the intracellular Ca2+ concentrations but also the Ca2+-dependent exocytotic processes. Of intracellular organelles that release Ca2+ in response to IP3, secretory granules stand out as the most prominent organelle and are responsible for the majority of IP3-dependent Ca2+ releases in the cytoplasm of chromaffin cells. Bovine chromaffin granules were the first granules that demonstrated the IP3-mediated Ca2+ release as well as the presence of the IP3 receptor (IP3R) in granule membranes. Secretory granules contain all three (type 1, 2, and 3) IP3R isoforms, and 58–69% of total cellular IP3R isoforms are expressed in bovine chromaffin granules. Moreover, secretory granules contain large amounts (2–4 mM) of chromogranins
and secretogranins; chromogranins A and B, and secretogranin II being the major species. Chromogranins A and B, and secretogranin
II are high-capacity, low-affinity Ca2+ binding proteins, binding 30–93 mol of Ca2+/mol of protein with dissociation constants of 1.5–4.0 mM. Due to this high Ca2+ storage properties of chromogranins secretory granules contain ~40 mM Ca2+. Furthermore, chromogranins A and B directly interact with the IP3Rs and modulate the IP3R/Ca2+ channels, i.e., increasing the open probability and the mean open time of the channels 8- to 16-fold and 9- to 42-fold, respectively.
Coupled chromogranins change the IP3R/Ca2+ channels to a more ordered, release-ready state, whereby making the IP3R/Ca2+ channels significantly more sensitive to IP3. 相似文献
997.
Sang Hyon Kim Mark Spensley Seung Kook Choi Cristiane P. G. Calixto Ali F. Pendle Olga Koroleva Peter J. Shaw John W. S. Brown 《Nucleic acids research》2010,38(9):3054-3067
Small nucleolar RNAs (snoRNAs) and small Cajal body-specific RNAs (scaRNAs) are non-coding RNAs whose main function in eukaryotes is to guide the modification of nucleotides in ribosomal and spliceosomal small nuclear RNAs, respectively. Full-length sequences of Arabidopsis snoRNAs and scaRNAs have been obtained from cDNA libraries of capped and uncapped small RNAs using RNA from isolated nucleoli from Arabidopsis cell cultures. We have identified 31 novel snoRNA genes (9 box C/D and 22 box H/ACA) and 15 new variants of previously described snoRNAs. Three related capped snoRNAs with a distinct gene organization and structure were identified as orthologues of animal U13snoRNAs. In addition, eight of the novel genes had no complementarity to rRNAs or snRNAs and are therefore putative orphan snoRNAs potentially reflecting wider functions for these RNAs. The nucleolar localization of a number of the snoRNAs and the localization to nuclear bodies of two putative scaRNAs was confirmed by in situ hybridization. The majority of the novel snoRNA genes were found in new gene clusters or as part of previously described clusters. These results expand the repertoire of Arabidopsis snoRNAs to 188 snoRNA genes with 294 gene variants. 相似文献
998.
The inulinase gene (INU1) from Kluyveromyces marxianus NCYC2887 strain was overexpressed by using GAL10 promotor in a △gal80 strain of Saccharomyces cerevisiae. The inulinase gene lacking the original signal sequence was fused in-frame to mating factor alpha signal sequence for secretory expression. Use of the △gal80 strain allowed the galactose-free induction of inulinase expression using a glucose-only medium. Shake flask cultivation in YPD medium produced 34.6 U/ml of the recombinant inulinase, which was approximately 13-fold higher than that produced by K. marxianus NCYC2887. It was found that the use of the △gal80 strain improved the expression of inulinase in the recombinant S. cerevisiae in both the aerobic and the anaerobic condition by about 2.9- and 1.7-fold, respectively. 5 L fed-batch fermentation using YPD medium was performed under aerobic condition with glucose feeding, which resulted in the inulinase production of 31.7 U/ml at OD600 of 67. Ethanol fermentation of dried powder of Jerusalem artichoke, an inulin-rich biomass, was also performed using the recombinant S. cerevisiae expressing INU1 and K. marxianus NCYC2887. Fermentation in a 5L scale fermentor was carried out at an aeration rate of 0.2 vvm, an agitation rate of 300 rpm, and the pH was controlled at 5.0. The temperature was maintained at 30degrees C and 37degrees C, respectively, for the recombinant S. cerevisiae and K. marxianus. The maximum productivities of ethanol were 59.0 and 53.5 g/L, respectively. 相似文献
999.
Background
Enzyme replacement therapy (ERT) with α-galactosidase A (α-Gal A) is currently the most effective therapeutic strategy for patients with Fabry disease, a lysosomal storage disease. However, ERT has limitations of a short half-life, requirement for frequent administration, and limited efficacy for patients with renal failure. Therefore, we investigated the efficacy of recombinant adeno-associated virus (rAAV) vector-mediated gene therapy for a Fabry disease mouse model and compared it with that of ERT.Methods
A pseudotyped rAAV2/8 vector encoding α-Gal A cDNA (rAAV2/8-hAGA) was prepared and injected into 18-week-old male Fabry mice through the tail vein. The α-Gal A expression level and globotriaosylceramide (Gb3) levels in the Fabry mice were examined and compared with Fabry mice with ERT. Immunohistochemical and ultrastructural studies were conducted.Results
Treatment of Fabry mice with rAAV2/8-hAGA resulted in the clearance of accumulated Gb3 in tissues such as liver, spleen, kidney, heart, and brain with concomitant elevation of α-Gal A enzyme activity. Enzyme activity was elevated for up to 60 weeks. In addition, expression of the α-Gal A protein was identified in the presence of rAAV2/8-hAGA at 6, 12, and 24 weeks after treatment. α-Gal A activity was significantly higher in the mice treated with rAAV2/8-hAGA than in Fabry mice that received ERT. Along with higher α-Gal A activity in the kidney of the Fabry mice treated with gene therapy, immunohistochemical studies showed more α-Gal A expression in the proximal tubules and glomerulus, and less Gb3 deposition in Fabry mice treated with this gene therapy than in mice given ERT. The α-gal A gene transfer significantly reduced the accumulation of Gb3 in the tubules and podocytes of the kidney. Electron microscopic analysis of the kidneys of Fabry mice also showed that gene therapy was more effective than ERT.Conclusions
The rAAV2/8-hAGA mediated α-Gal A gene therapy provided improved efficiency over ERT in the Fabry disease mouse model. Furthermore, rAAV2/8-hAGA-mediated expression showed a greater effect in the kidney than ERT.1000.
Choong Hyun Lee Ki-Yeon Yoo Ok Kyu Park Jung Hoon Choi Il-Jun Kang Eunjoo Bae Sung Koo Kim In Koo Hwang Moo-Ho Won 《Molecules and cells》2010,29(4):373-378
Phosphorylated extracellular signal-regulated kinase (pERK) mediates neuronal synaptic plasticity, long-term potentiation,
and learning and memory in the hippocampus. In this study, we examined pERK1/2 immunoreactivity and its protein level in the
gerbil hippocampus at various ages. In the postnatal month 1 (PM 1) group, very weak pERK1/2 immunoreactivity was detected
in the hippocampus. In the CA1 region, pERK1/2 immunoreactivity was considerably increased in the stratum pyramidale in the
PM 6 group. Thereafter, pERK1/2 immunoreactivity was decreased. In the CA2/3 region, pERK1/2 immunoreactivity increased in
an age-dependent manner until PM 12. Thereafter, numbers of pERK1/2-immunoreactive neurons were decreased. However, in the
mossy fiber zone, pERK1/2 immunostaining became stronger with age. In the dentate gyrus, a few pERK1/2-immunoreactive cells
were observed until PM 12. In the PM 18 and 24 groups, numbers of pERK1/2-immunoreactive cells were increased, especially
in the polymorphic layer. In Western blot analysis, pERK1/2 level in the gerbil hippocampus was increased with age. These
results indicate that total pERK1/2 levels are increased in the hippocampus with age. However pERK1/2 immunoreactivity in
subregions of the gerbil hippocampus was changed with different pattern during normal aging. 相似文献