• Journal of the korean academy of Pediatric Dentistry
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• v.28 no.2
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• pp.219-227
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• 2001

Little is known about processing mechanism of pain sensation of the oral cavity at the 1st synapse of trigeminal sensory nuclei. Serial ultrathin sections of tooth pulp afferent terminals, identified by the transganglionic transport of 1% wheatgerm agglutinin conjugated horseradish peroxidase, were investigated with electron microscope. Quantitative ultrastructural analysis was performed on digitizing tablet connected to Macintoshi personal computer (software; NIH Image 1.60, NIH, Bethesda, MD). Labeled boutons could be classified into two types by the shapes of containing vesicles : S bouton, which contained mainly spherical vesicles (Dia. 45-55 nm) and few large dense cored vesicles (Dia, 80-120nm), and LDCV bouton, which contained spherical vesicles as well as large number of large dense cored vesicles. Most of the parameters on the ultrastructural characteristic and synaptic organization of labeled boutons were similar between S and LDCV boutons, except shapes of containing vesicles. Majority of the labeled boutons showed simple synaptic arrangement. The labeled boutons were frequency presynaptic to dendritic spine, and to a lesser extent, dendritic shaft. They rarely synapsed with soma and adjacent proximal dendrite. A small proportion of labeled boutons made synaptic contacts with presynaptic, pleomorphic vesicles containing endings and synaptic triad. Morphometric parameters of labeled boutons including volume and surface area, total apposed area, mitochondrial volume, active zone area, vesicle number and density showed wide variation and these were not significantly different between S and LDCV boutons. The present study revealed characteristic features on ultrastructure and synaptic connection of pulpal afferents which may involved in transmission of oral pain sensation.

• Journal of Life Science
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• v.28 no.12
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• pp.1416-1423
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• 2018

With strong biotin binding affinity ($K_D=10^{-14}M$), the tetrameric feature of streptavidin could be used to increase the antigen binding activity of a camel heavy chain (VHH) antibody through their fusion, here stained with biotinylated horseradish peroxidase and subsequent immunoassays ELISA and Western blot analysis. For this application, we cloned the streptavidin gene amplified from the Streptomyces avidinii chromosome by PCR, and this was fused to the gene of the 8B9 VHH antibody which is specific to green fluorescent protein (GFP) antigens. To express a soluble fusion protein in Escherichia coli, we used the pUC119 plasmid-based expression system which uses the lacZ promoter for induction by IPTG, the pelB leader sequence at the N-terminus for secretion into the periplasmic space, and six polyhistidine tags at the C-terminus for purification of the expressed proteins using an $Ni^+$-NTA-agarose column. Although streptavidin is toxic to E. coli because of its strong biotin binding property, this soluble fusion protein was expressed successfully. In SDS-PAGE, the size of the purified fusion protein was 122.4 kDa in its native condition and 30.6 kDa once denatured by boiling, suggesting the tetramerization of the monomeric subunit by non-covalent association through the streptavidin moiety fusing to the 8B9 VHH antibody. In addition, this fusion protein showed biotin binding activity similar to streptavidin as well as GFP antigen binding activity through both ELISA and Western blot analysis. In conclusion, the protein resulting from the fusion of an 8B9 VHH antibody with streptavidin was successfully expressed and purified as a soluble tetramer in E. coli; it showed both biotin and GFP antigen binding activity suggesting the possible production of a tetrameric and bifunctional VHH antibody.

• Applied Microscopy
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• v.23 no.1
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• pp.1-14
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• 1993

The present study was carried out to analyze the evidence of membrane recycling, and the regulation of cellular transport by dynamic changes in apical membrane area that functionally interacts with the number of cytoplasmic vesicles. Under scanning electron micrographs, turtle bladder mucosa contain three main type of cells; granular cells and carbonic anhydrase (CA)-rich cells, deviding into a and b type of epithelial cell. The granular cell is the majority cell type of the mucosa comprising 80% of the total cell number. The remaining 20% of the cells are characteristically rich in carbonic anhydrase. Uptake of HRP was detected in the most vacuoles or tubulovesicles in both type of CA-rich cells in the turtle bladder, indicating that the part of plasma membrane was internalized in the apical cytoplasmic vacuoles. It seems quite likely that CA-rich cells possess intracellular vesicles carrying proton pumps which are recycling back to the apical plasma membrane. In turtle bladder, the granular cells actively secrete large quantities of mucin and other proteins by an exocytotic mechanism in an apparently constitutive fashion. The possibility that bladder epithelial cells secrete mucin via a regulated secretory pathway has not been rigorously examined and much is still to be determined about these issues from this cell type.

• Korean Journal of Environmental Agriculture
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• v.3 no.2
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• pp.37-42
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• 1984

The effect on the activities of peroxidase and lipoxygenase by chemicals was determined. Peroxidase activities of horseradish and apple were inhibited strongly by maneb and comparativelyby mancozeb, zineb. The inhibitory rate of enzyme activity was ranged in $12.5{\sim}34.5%$ by maneb, $11.0{\sim}30.5%$ by mancozeb, and $9.7{\sim}27.5%$ by zineb in the level of $10{\sim}30\;ppm$ at the final concentration. No signification was in the peroxidase activity of apple during ripening between control and plot, treated with 500 ppm mancozeb. On the activity of purified soybean lipoygenase, zineb inhibited it more strongly than carbofuran or phosphamidon, but maneb did not have inhibitory effect on that. The inhibition ranges of $14.0{\sim}40.0%$ by zineb, $6.5{\sim}20.0%$ by carbofuran and $4.5{\sim}13.0%$ by phosphamidon were shown in the final pesticide concentration of $10{\sim}30$ ppm. But in $100{\sim}200$ ppm, lipoxygenase activity was almost inhibited by carbofuran and phosphamidon.

• Korean Journal of Food Science and Technology
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• v.36 no.6
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• pp.959-967
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• 2004

To screen probiotic lactic acid bacteria with potent adhesive property on human colonic mucosa, colonic mucin-binding assay was introduced. This colonic mucin-binding assay actually measures the binding activity of surface lectin-like protein (SLP) on colonic mucin, and the optimal conditions were examined. The optimal pH for colonic mucin coating on plate wells was 4.8, and ${\times}24,000$ diluted solution of commercially available horseradish peroxidase (HRP) conjugated streptoavidin yielded good results, for rapid screening, $5.0\;{\mu}g/mL$ of biotinylated SLP from lactic acid bacteria was optimal, and optimal scintillation time of 3,3',5,5'-tetramethyl benzidine (TMB) was 10 min. These conditions were useful for both rapid selection and quantitative analysis of lactic acid bacteria that have high adhesion property to human intestinal tract. Among 50 strains of lactic acid bacteria, including 32 type culture strains and 18 isolated strains from infant feces, Lactobacillus species FSB-1 isolated from kimchi showed the highest binding activity to colonic mucin. From taxonomical viewpoints based on morphological study, physico-biochemical study, partial 16S rDNA seguencing, and phylogenetic analysis, L. species FSB-1 was identified as Lactobacillus brevis.

• Korean Journal of Food Science and Technology
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• v.46 no.6
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• pp.689-693
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• 2014

Flounder sikhe is one of the traditional Korean fermented foods. Because honey, fucoidan, propolis, aronia berry, and horseradish are well known to have natural antimicrobial properties, we investigated their antimicrobial effects on improving the quality of flounder sikhe. Ethanol extracts of propolis inhibited the growth of Bacillus cereus, Staphylococcus aureus, and Listeria monocytogenes. The extracts of aronia berry showed similar effects against Salmonella Typhimurium. We also assessed the total polyphenol and total flavonoid content and 2,2-diphenyl-2-picrythydrazyl (DPPH) radical scavenging activity in the extracts obtained from these natural materials. The contents of polyphenols and flavonoids were the highest in propolis, which also showed a high DPPH radical scavenging activity. Therefore, addition of propolis 0.5% to flounder sikhe showed optimal improvement, with a storage stability of 15 days at pH 5.39, volatile basic nitrogen (VBN) content at 24.73 mg/100 g, and total bacterial count at $1.81{\times}10$ colony forming unit (CFU)/g.

• Korean Journal of Environmental Agriculture
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• v.18 no.3
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• pp.292-297
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• 1999

This study was performed to estimate the possibility of use of plant materials as catalytic agents fur the decontamination of waste waters contaminated with organic pollutants by using 2,4-dichlorophenol(2,4-DCP) as a model pollutant. Plant materials containing high peroxidase activity were selected as catalysts for the removal of 2,4-DCP. Peroxidase activity, which plant materials were containing, was measured, and the greatest peroxidase activity was observed in shepherd's purse, followed by turnip, sweet potato, Chinese cabbage and white radish. The peroxidase activity in shepherd's purse was four times higher than that of horseradish purchased in U.S.A. Using shepherd' s purse and turnip, it was investigated the effect of various factors on the decontamination of 2,4-DCP through oxidative coupling. The removal of 2,4-DCP was extremely fast, and a maximal removal could be achieved within 3 min for shepherd' s purse and 15min for turnip. The pH range was from 3.0 to 8.0 and the amount of $H_2O_2$ added was 9mM when maximal removal was achieved(over 90%). No increasing removal of 2,4-DCP was observed due to increasing the amount of $H_2O_2$ added (over 9mM). The initial concentration affected the transformation of 2,4-DCP incubated with plant materials. When turnip was used as catalytic agent, it was observed decreasing transformation of 2,4-DCP due to increasing initial concentration.

• Applied Microscopy
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• v.38 no.4
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• pp.375-382
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• 2008

In order to investigate shape of synaptic vesicles of the tooth pulp afferent boutons and their presynaptic endings (p-endings), and the neuroactive substance of the p-endings in the trigeminal nucleus principalis, rat incisor tooth pulp afferents were labeled by the horseradish peroxidase (HRP) and quantitative ultrastructural analysis and postembedding immunogold labeling were performed. Labeled tooth pulp afferent boutons contained clear, spherical synaptic vesicles (diameter: $45{\sim}55\;nm$) and occasionally dense core vesicles(diameter: $80{\sim}120\;nm$). They formed symmetrical synapses with unlabeled axon terminals (p-endings) containing pleomorphic synaptic vesicles. The ratio of short to long diameter (form factor) of synaptic vesicles of pulp afferent boutons was 0.6 to 0.99, whereas that of p-endings was 0.25 to 0.99. In addition, most of the p-endings showed GABA-like immunoreactivity. These results indicate that the shape of synaptic vesicles is quite different between the tooth pulp afferent boutons and p-endings, and the p-endings may contain GABA as a neuroactive substance in the trigeminal nucleus principalis.

• Korean Journal of Food Science and Technology
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• v.35 no.3
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• pp.366-372
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• 2003

Enzyme-linked immunosorbent assay (ELISA) for assaying the 5-enolpyruvyshikimate-3-phosphate synthase from Agribacterium sp. CP4 (CP4 EPSPS) in genetically modified soybeans was developed. Polyclonal and monoclonal antibodies (Pab, Mab) specific to the CP4 EPSPS were produced. When using the Pab, the detection limit of sandwich ELISA toward CP4 EPSPS (0.03 ${\mu}g/mL$) was better than that of competitive indirect ELISA(ciELISA) (1 ${\mu}g/mL$). It was found that 2 of 3 monoclonal antibodies, Mab1 and Mab2, recognized the same antigenic determinant on CP4 EPSPS, but Mab3 recognized different antigenic determinant when competitive ELISA was performed using the Mabs. On the other hand, when the sensitivity of sandwich ELISA using combination of Pab and/or Mabs was determined, the sandiwich ELISA using Mab2 as a capture antibody and Pab-HRP as a secondary antibody showed the lowest detection limit of CP4 EPSPS (0.02 ${\mu}g/mL$). The sandwich ELISA developed in this study could be applied to detect glyphosate-tolerant soybeans.

• Molecules and Cells
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• v.24 no.2
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• pp.276-282
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• 2007

Protein phosphorylation is one of the major mechanisms by which eukaryotic cells transduce extracellular signals into intracellular responses. Calcium/calmodulin ($Ca^{2+}/CaM$)-dependent protein phosphorylation has been implicated in various cellular processes, yet little is known about $Ca^{2+}/CaM$-dependent protein kinases (CaMKs) in plants. From an Arabidopsis expression library screen using a horseradish peroxidase-conjugated soybean calmodulin isoform (SCaM-1) as a probe, we isolated a full-length cDNA clone that encodes AtCK (Arabidopsis thaliana calcium/calmodulin-dependent protein kinase). The predicted structure of AtCK contains a serine/threonine protein kinase catalytic domain followed by a putative calmodulin-binding domain and a putative $Ca^{2+}$-binding domain. Recombinant AtCK was expressed in E. coli and bound to calmodulin in a $Ca^{2+}$-dependent manner. The ability of CaM to bind to AtCK was confirmed by gel mobility shift and competition assays. AtCK exhibited its highest levels of autophosphorylation in the presence of 3 mM $Mn^{2+}$. The phosphorylation of myelin basic protein (MBP) by AtCK was enhanced when AtCK was under the control of calcium-bound CaM, as previously observed for other $Ca^{2+}/CaM$-dependent protein kinases. In contrast to maize and tobacco CCaMKs (calcium and $Ca^{2+}/CaM$-dependent protein kinase), increasing the concentration of calmodulin to more than $3{\mu}M$ suppressed the phosphorylation activity of AtCK. Taken together our results indicate that AtCK is a novel Arabidopsis $Ca^{2+}/CaM$-dependent protein kinase which is presumably involved in CaM-mediated signaling.