• Journal of Plant Biotechnology
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• v.43 no.2
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• pp.231-239
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• 2016

To study the transcript levels of epigenetically regulated genes in tobacco, we have developed a transgenic line OX1 overexpressing NtROS2a gene encoding cytosine DNA demethylation and a RNAi plant line RNAi13. It has been reported that salt- and $H_2O_2$-stress tolerance of these transgenic lines are enhanced with various phenotypic characters (Lee et al. 2015). In this paper, we conducted microarray analysis with Agilent Tobacco 4 x 44K oligo chip by using overexpression line OX1, RNAi plant line RNAi 13, and wild type plant WT. Differentially expressed genes (DEGs) related to metabolism, nutrient supply, and various stressed were up-regulated by approximately 1.5- to 80- fold. DEGs related to co-enzymes, metabolism, and methylation functional genes were down-regulated by approximately 0.03- to 0.7- fold. qRT-PCR analysis showed that the transcript levels of several candidate genes in OX1 and RNAi lines were significantly (p < 0.05) higher than those in WT, such as genes encoding KH domain-containing protein, MADS-box protein, and Zinc phosphodiesterase ELAC protein. On the other hand, several genes such as those encoding pentatricopeptide (PPR) repeat-containing protein, histone deacetylase HDAC3 protein, and protein kinase were decreased by approximately 0.4- to 1.0- fold. This study showed that NtROS2a gene encoding DNA glycosylase related to demethylation could regulate adaptive response of tobacco at transcriptional level.

• Biomolecules & Therapeutics
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• v.11 no.3
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• pp.157-162
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• 2003

The generation of secretory IgA antibodies(Abs) for specific immune protection of mucosal surfaces depends on stimulation of the mucosal immune system, but this is not effectively achieved by parenteral or even oral administration of most soluble antigens. Thus, to produce a possible vaccine antigen against urinary tract infections, the uropathogenic E. coli (UPEC) adhesin was genetically coupled to the ctxa2b gene and cloned into a pMAL-p2E expression vector. The chimeric construction of pMALfimHIctxa2b was then transformed into E. coli K-12 TB1 and its nucleotide sequence was verified. The chimeric protein was then purified by applying the affinity chromatography. The purified chimeric protein was confirmed by SDS-PAGE and western blotting using antibodies to the maltose binding protein (MBP) or the cholera toxin subunit B (CTXB), plus the N-terminal amino acid sequence was analyzed. The orderly-assembled chimeric protein was confirmed by a modified $G_{M1}$-ganglioside ELISA using antibodies to adhesin. The results indicate that the purified chimeric protein was an Adhesin/CTXA2B protein containing UPEC adhesin and the $G_{M1}$-ganglioside binding activity of CTXB. This study also demonstrate that peroral administration of this chimeric immunogen in mice elicited high level of secretory IgA and serum IgG Abs to the UPEC adhesin. The results suggest that the genetically linked CTXA2B acts as a useful mucosal adjuvant, and that the adhesin/CTXA2B chimeric protein might be a potential antigen for oral immunization against UPEC.

• The Korean Journal of Zoology
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• v.32 no.2
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• pp.153-162
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• 1989

Expedments were perfonned to examine the role of cAMP-dependent protein kinase (PK-A) and diacylglycerol-dependent protein kinase (PK-C) during the meiodc resumption and the first mitotic cell cycle of mouse embryogenesis. Mejoric GV oocytes and one-cell embryos derived from in vitro fertilization were cultured in vitro, and morphological changes in response to activators of PK-A and PK-C were examined. Treatments with a membrane-permeable cAMP analog, dbcAMP (0.1 mg/mi), phosphodiesterase inhibitor, IBMX (0.1 mM), biologically active phorbol ester, WA (10 nglmi), or a synthetic diacylglycerol, sn-diC8 inhibited resumption of melosis. Combination of PK-A and PK-C activator brought about furiher inhibition. On the contrary, dbcAMP (0.1 mg/mi), IBMX (0.2 mM), WA (10 nglml), and sn-diC8 (0.5 mM) did not inhibit pronucleus membrane breakdown (PNBD) when added S or G2 phase of cell cycle. However, activators of PK-C inhibited cleavage of one-cefl embryos. This result indicates that the action mechanism of PK-A and PK-C on dissolution of nuclear membrane in primary meiotic arrest oocytes may be different from that of mitotic one-cell embryos.

• Genomics & Informatics
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• v.9 no.2
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• pp.74-78
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• 2011

Numerous restraints and simplifications have been developed for methods that anticipate protein structure to reduce the colossal magnitude of possible conformational states. In this study, we investigated if globularity is a general characteristic of proteins and whether they can be applied as a valid constraint in protein structure simulations with approximated measurements (Gb-index). Unexpectedly, most of the proteins showed strong structural globularity (i.e., mode of approximately 76% similarity to the perfect globe) with only a few percent of proteins being outliers. Small proteins tended to be significantly non-globular ($R^2$=0.79) and the minimum Gb-index showed a logarithmic increase with the increase in protein size ($R^2$=0.62), strongly implying that the non-globular characteristics might be more acceptable for smaller proteins than larger ones. The strong perfect globe-like character and the relationship between small size and the loss of globular structure of a protein may imply that living organisms have mechanisms to aid folding into the globular structure to reduce irreversible aggregation. This also implies the possible mechanisms of diseases caused by protein aggregation, including some forms of trinucleotide repeat expansion-mediated diseases.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.1
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• pp.55-61
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• 2014

Forty-eight Pelibuey${\times}$Katahdin male intact lambs ($23.87{\pm}2.84$ kg) were used in an 84-d feeding trial, with six pens per treatment in a $2{\times}2$ factorial design arrangement. The aim of the study was to evaluate the interaction of two dietary energy levels (3.05 and 2.83 Mcal/kg ME) and two dietary protein levels (17.5% and 14.5%) on growth performance, dietary energetics and carcass traits. The dietary treatments used were: i) High protein-high energy (HP-HE); ii) High protein-low energy (HP-LE); iii) Low protein-high energy (LP-HE), and iv) Low protein-low energy (LP-LE). With a high-energy level, dry matter intake (DMI) values were 6.1% lower in the low-protein diets, while with low-energy, the DMI values did not differ between the dietary protein levels. Energy levels did not influence the final weight and average daily gain (ADG), but resulted in lower DMI values and higher gain efficiencies. No effects of protein level were detected on growth performance. The observed dietary net energy (NE) ratio and observed DMI were closer than expected in all treatments and were not affected by the different treatments. There was an interaction (p<0.03) between energy and protein level for kidney-pelvic and heart fat (KPH), KPH was higher in lambs fed high energy and high protein diet but not in high energy and low protein diet. The KPH was increased (20.2%, p = 0.01) in high-energy diets, while fat thickness was increased (21.7%, p = 0.02) in high-protein diets. Therefore, it is concluded that dietary energy levels play a more important role in feed efficiency than protein levels in finishing lambs with a high-energy diet (>2.80 Mcal/kg ME). Providing a level of protein above 14.5% does not improves growth-performance, dietary energetics or carcass dressing percentage.

• Biomedical Science Letters
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• v.18 no.2
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• pp.96-103
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• 2012

Kv 4.2 ion channel protein has an ability to open at subthreshold membrane potentials and to recover quickly from inactivation. That is very important for neuronal signal transmission in vertebrate brain. In order to purify Kv 4.2 protein, the novel purification methods were experimented. The purification procedure utilized chromatography on DE-52 ion exchange column and affinity chromatography on a WGA-Sepharose 4B, and Kv 4.2 affinity column chromatography. It was found that 0.5% (wt./vol.) Triton X-100 detergent in lysis buffer worked well for Kv 4.2 protein solubilization from rat brain membrane. Protein quantitative determination was conducted by BCA method at 562 nm for each purification step to avoid determination interference of protein at 280 nm by detergent. The confirmation of Kv 4.2 existence and amount is performed using by SDS-PAGE/immunoblotting or 96-well dot blotting. The Kv 4.2 without interacting protein that contains carbohydrate, was purified from novel biochemical 3-steps purification method for further research.

• Korean Journal of Agricultural Science
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• v.37 no.2
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• pp.245-248
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• 2010

This study was conducted to investigate effect of changes in target fat and protein contents in milk on feed cost using a simulation modeling approach based on the 2001 dairy NRC. Two simulations were done; simulation I had a limitation (up to 20%), but simulation II had no limitation for the use of cottonseed hull in a diet. Using commonly used feed ingredients in Korea, we formulated least cost diets that meet nutrient requirement of a lactating dairy cow producing 36 kg of milk with combinations of 0.1% decrease or 0.1% increase in target milk fat or protein, respectively, from the national average milk fat (4.0%) and milk protein (3.1%). The contents of alfalfa and corn in a least-cost diet were decreased and those of tall fescue, whole cottonseed and rapeseed meal were increased with decreasing fat and/or increasing protein in milk. Scenarios that decreased target milk fat percentage from 4.0% to 3.9% reduced feed cost by 2 won per kg. Due to decrease in feed intake, daily feed cost was even more reduced (136 won per head) by decreasing target milk fat percentage. Increase in target milk protein percentage from 3.1% to 3.2% reduced feed cost by 6 won per kg. Among scenarios simulated, the least feed cost was obtained in scenario aimed for 3.9% fat and 3.2% of protein in milk. We conclude that a feeding practice for increasing milk protein percentage does not directly increase feed cost. In addition, feeding practices that increase protein content in milk is expected to improve economic life-span and reproductive performance of dairy cows.

• BMB Reports
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• v.33 no.2
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• pp.103-111
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• 2000

Phosphorylation of the eukaryotic elongation factor 2 (eEF-2) blocks the elongation step of translation and stops overall protein synthesis. Although the overall rate of protein synthesis in mitosis reduces to 20% of that in S phase, it is unclear how the protein translation procedure is regulated during the cell cycle, especially in the stage of peptide elongation. To delineate the regulation of the elongation step through eEF-2 function, the changes in phosphorylation of eEF-2, and in activity of corresponding $Ca^{2+}$/calmodulin (CaM)-dependent protein kinase III (CaMK-III) during the cell cycle of NIH 3T3 cells, were determined. The in vivo level of phosphorylated eEF-2 showed an 80% and 40% increase in the cells arrested at G1 and M, respectively. The activity of CaMK-III also changed in a similar pattern, more than a 2-fold increase when arrested at G1 and M. The activity change of the kinase during one turn of the cell cycle also demonstrated the activation at G1 and M phases. The activity change of cAMP-dependent protein kinase (PKA) was reciprocal to that of CaMK-III. These results indicated: (1) the activity of CaMK-III was cell cycle-dependent and (2) the level of eEF-2 phosphorylation followed the kinase activity change. Therefore, the elongation step of protein synthesis might be cell cycle dependently regulated.

• YAKHAK HOEJI
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• v.45 no.3
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• pp.287-292
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• 2001

To investigate whether phospholipase $A_2$pathway is involved in histamine release of rat peritoneal mast cells, we measured histamine release in the presence of various enzyme inhibitors involved in eicosanoid pathway, such as phospholipase $A_2$, cyclooxygenase and lipoxygenase. Phospholipase $A_2$inhibitors, manoalide and OPC, significantly inhibited histamine release induced by 100 $\mu$M ATP and 1$\mu$g/ml compound 48/80. Cyclooxygenase inhibitors, ibuprofen and indomethacin, significantly inhibited ATP-induced histamine release and lipoxygenase inhibitors, baicalein and caffeic acid, also significantly inhibited. To investigate the involvement of protein kinase in ATP- and compound 48/80-induced histamine release, we observed effects of protein kinase inhibitors on histamine release. Bisindolmaleimide (protein kinase C antagonist) dose-dependently inhibited both ATP and compound 48/80-induced histamine release. Tyrosine kinase inhibitors (methyl 2,5-dihydroxy cinnamate and genistein) dose-dependently inhibited ATP and compound 48/80-induced histamine release. Protein kinase C and tyrosine kinase seem to be involved in histamine release induced by ATP and compound 48/80. These results suggest that phospholipase $A_2$pathway as well as protein kinase C and tyrosine kinase are involved in histamine release of rat peritoneal mast cells by ATP and compound 48/80.

• Journal of Nutrition and Health
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• v.22 no.6
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• pp.497-506
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• 1989

To investigate the effect of levels of dietary protein and age on bone metabolism 40% and 5% casein were fed to the rats of 2 & 13 months of age for 12 weeks. High protein groups showed higher bone weight and Ca content than low protein groups and urinary Ca loss was increased in high protein groups but the difference disappeared gradually. A significant increase in urinary hydroxyproline excretion was noted in high protein groups of both age. Another short term study was undertaken to study if the above effect was related with renal function or PTH. Extremely high and low protein diets(60%, 6%) were fed to the rats of different ages(6wks, 6mos.) for 2 weeks, Urinary Ca excretion was significantly increased in high protein groups of young and aged rats and GFR was increased as well. There was no difference in serum iPTH levels between low and high protein groups, but it was elevated in aged rats. Alkaline phosphatase activity was higher in young rats, reflecting faster bone formation. The observed hypercalciuria in high protein groups, especially in aged rats, seems to be related to higher GFR, and PTH dose not appear to be a major mediator.