• Reproductive and Developmental Biology
• /
• v.39 no.4
• /
• pp.105-115
• /
• 2015

Toll-like receptor 7 (TLR7) is critical for the triggering of innate immune response by recognizing the conserved molecular patterns of single-stranded RNA (ssRNA) viruses and mediated antigenic adaptive immunity. To understand how TLR7 distinguish pathogen-derived molecular patterns from the host self, it is essential to be able to identify TLR7 receptor interaction interfaces, such as active sites or R848-agonist binding sites. The functional interfaces of TLR7 can serve as targets for structure-based drug design in studying the TLR7 receptor's structure-function relationship. In contrast to mammalian TLR7, chicken TLR7 (chTLR7) is unknown for its important biological function. Therefore, it has been targeted to mediate contrasting evolutionary patterns of positive selection into non-synonymous SNPs across eleven species using TLR7 conservation patterns (evolutionary conserved and class-specific trace residues), where protein sequence differences to the TLR7 receptors of interest record mutation that have passed positive section across the species. In this study, we characterized the Lys609 residue on chTLR7-ECD homodimer interfaces to reflect the current tendency of evolving positive selection to be transfer into a stabilization direction of the R848-agonist/chTLR7-ECDs complex under the phylogenetically variable position across species and we suggest a potential indicator for contrasting evolutionary patterns of both the species TLR-ECDs.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2014.02a
• /
• pp.126-126
• /
• 2014

Label-free biomolecular assay based localized surface plasmon resonance (LSPR) of noble metal nanoparticles enables simple and rapid detection with the use of simple equipment. Nanosized metal nanoparticles exhibit a strong absorption band when the incident light frequency is resonant with the collective oscillation of the electrons, which is known as the LSPR. Here we demonstrate localized surface plasmon resonance (LSPR) substrates such as plasmonic Au nanodisks fabricated by a nanoimprinting process and gold nanorod-immobilized surfaces and their applications to highly sensitive and/or label-free biosensing. To increase detection sensitivity various bioreceptors weree designed. A single chain variable fragment (scFv) was used as a receptor to bind C-reactive protein (CRP). The results of this effort showed that CRP in human serum could be quantitatively detected lower than 1 ng/ml. Aptamers, which were immobilized on gold nanorods, were used to detect mycotoxins. The specific binding of ochratoxin A (OTA) to the aptamer was monitored by the longitudinal wavelength shift of LSPR peak in the UV-Vis spectra resulting from the changes of local refractive index near the GNR surface induced by accumulation of OTA and G-quadruplex structure formation of the aptamer. According to our results, OTA could be quantitatively detected lower than 1 nM level. Additionally, aptamer-functionalized GNR substrate was quite robust and can be regenerated many times by rinsing at 70 OC to remove bound target. During seven times of washing steps, the developed OTA sensing system could be reusable. Moreover, the proposed biosensor exhibited selectivity over other mycotoxins with an excellent recovery for detection in grinded corn samples, suggesting that the proposed LSPR based aptasensor plays an important role in label-free detection of mycotoxins.

• Proceedings of the Korean Vacuum Society Conference
• /
• 2011.02a
• /
• pp.252-253
• /
• 2011

ZnO-based thin film transistors (TFTs) are of great interest for application in next generation flat panel displays. Most research has been based on amorphous indium-gallium-zinc-oxide (IGZO) TFTs, rather than single binary oxides, such as ZnO, due to the reproducibility, uniformity, and surface smoothness of the IGZO active channel layer. However, recently, intrinsic ZnO-TFTs have been investigated, and TFT- arrayss have been demonstrated as prototypes of flat-panel displays and electronic circuits. However, ZnO thin films have some significant problems for application as an active channel layer of TFTs; it was easy to change the electrical properties of the i-ZnO thin films under external conditions. The variable electrical properties lead to unstable TFTs device characteristics under bias stress and/or temperature. In order to obtain higher performance and more stable ZnO-based TFTs, HZO thin film was used as an active channel layer. It was expected that HZO-TFTs would have more stable electrical characteristics under gate bias stress conditions because the binding energy of Hf-O is greater than that of Zn-O. For deposition of HZO thin films, Hf would be substituted with Zn, and then Hf could be suppressed to generate oxygen vacancies. In this study, the fabrication of the oxide-based TFTs with HZO active channel layer was reported with excellent stability. Application of HZO thin films as an active channel layer improved the TFT device performance and bias stability, as compared to i-ZnO TFTs. The excellent negative bias temperature stress (NBTS) stability of the device was analyzed using the HZO and i-ZnO TFTs transfer curves acquired at a high temperature (473 K).

• Archives of Pharmacal Research
• /
• v.29 no.8
• /
• pp.677-684
• /
• 2006

Morphine has been used widely on the treatment of many types of chronic pain. However the development of tolerance to and dependence on morphine by repeat application is a major problem in pain therapy. The purpose of the present study was to investigate whether combined administration of nalbuphine with morphine affects the development of tolerance to and dependence on morphine. We hypothesize that the use of nalbuphine, ${\kappa}-agonist$ may prove to be useful adjunct therapy to prevent morphine-induced undesirable effects in the management of some forms of chronic pain. Morphine (10 mg/kg) was injected to rats intraperitoneally for 5 day. The variable dose of nalbuphine (0.1, 1.0 and 5.0 mg/kg) was administered (i.p.) in combination with morphine injection. The development of morphine tolerance was assessed by measuring the antinociceptive effect with the Randall-Selitto apparatus. The development of dependence on morphine was determined by the scoring the precipitated withdrawal signs for 30 min after injection of naloxone (10 mg/kg, i.p.). Nalbuphine did not attenuate antinociceptive effect of morphine in rats. Interestingly, combined administration of morphine with nalbuphine (10:1) significantly attenuated the development of dependence on morphine. The elevation of $[^3H]MK-801$ binding in frontal cortex, dentate gyrus, and cerebellum after chronic morphine infusion was suppressed by the coadministration of nalbuphine. In addition, the elevation of NR1 expression by morphine was decreased by the coadministration of nalbuphine in rat cortex. These results suggest that the coadministration of nalbuphine with morphine in chronic pain treatment can be one of therapies to reduce the development of tolerance to and dependence on morphine.

• Journal of Korean Medicine for Obesity Research
• /
• v.20 no.1
• /
• pp.1-9
• /
• 2020

Objectives: Artemisinin and its derivatives extracted from Artemisia annua, a Chinese herbal medicine, have variable biological effects due to structural differences. Up to date, the anti-obesity effect of dihydroartemisinin (DHA), a derivative of artemisinin, is unknown. The purpose of this study was to investigate the anti-adipogenic and lipolytic effects of DHA on 3T3-L1 preadipocytes. Methods: Oil Red O staining and AdipoRed assay were used to measure lipid accumulation and triglyceride (TG) content in 3T3-L1 cells, respectively. Cell count analysis was used to determine the cytotoxicity of 3T3-L1 cells. Western blot and real-time reverse transcription polymerase chain reaction analyses were used to analyze the expression of protein and mRNA in 3T3-L1 cells, respectively. Results: DHA at 5 μM markedly inhibited lipid accumulation and reduced TG content in differentiating 3T3-L1 cells with no cytotoxicity. Furthermore, DHA at 5 μM inhibited the expression of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), fatty acid synthase (FAS), and perilipin A as well as the phosphorylation of signal transducer and activator of transcription-3 (STAT-3) in differentiating 3T3-L1 cells. Moreover, while DHA at 5 μM had no effect on the mRNA expression of adiponectin, it strongly suppressed that of leptin in differentiating 3T3-L1 cells. However, DHA at 5 μM had no lipolytic effect on differentiated 3T3-L1 cells, as assessed by no enhancement of glycerol release. Conclusions: These results demonstrate that DHA at 5 μM has a strong anti-adipogenic effect on differentiating 3T3-L1 cells through the reduced expression and phosphorylation of C/EBP-α, PPAR-γ, FAS, perilipin A, and STAT-3.

• Journal of Microbiology and Biotechnology
• /
• v.24 no.3
• /
• pp.408-420
• /
• 2014

Yeast surface-displayed antibody libraries provide an efficient and quantitative screening resource for given antigens, but suffer from typically modest library sizes owing to low yeast transformation efficiency. Yeast mating is an attractive method for overcoming the limit of yeast transformation to construct a large, combinatorial antibody library, but the optimal conditions have not been reported. Here, we report a large synthetic human Fab (antigen binding fragment) yeast surface-displayed library generated by stepwise optimization of yeast mating conditions. We first constructed HC (heavy chain) and LC (light chain) libraries, where all of the six CDRs (complementarity-determining regions) of the variable domains were diversified mimicking the human germline antibody repertoires by degenerate codons, onto single frameworks of VH3-23 and $V{\kappa}1$-16 germline sequences, in two haploid cells of opposite mating types. Yeast mating conditions were optimized in the order of cell density, media pH, and cell growth phase, yielding a mating efficiency of ~58% between the two haploid cells carrying HC and LC libraries. We constructed two combinatorial Fab libraries with CDR-H3 of 9 or 11 residues in length with colony diversities of more than $10^9$ by one round of yeast mating between the two haploid HC and LC libraries, with modest diversity sizes of ${\sim}10^7$. The synthetic human Fab yeast-displayed libraries exhibited relative amino acid compositions in each position of the six CDRs that were very similar to those of the designed repertoires, suggesting that they are a promising source for human Fab antibody screening.

• Journal of Veterinary Science
• /
• v.22 no.4
• /
• pp.45.1-45.13
• /
• 2021

Background: Peste des petits ruminants (PPR) is an infectious disease caused by the peste des petits ruminants virus (PPRV) that mainly produces respiratory symptoms in affected animals, resulting in great losses in the world's agriculture industry every year. Single-domain variable heavy chain (VHH) antibody fragments, also referred to as nanobodies, have high expression yields and other advantages including ease of purification and high solubility. Objectives: The purpose of this study is to obtain a single-domain antibody with good reactivity and high specificity against PPRV. Methods: A VHH cDNA library was established by immunizing camels with PPRV vaccine, and the capacity and diversity of the library were examined. Four PPRV VHHs were selected, and the biological activity and antigen-binding capacity of the four VHHs were identified by western blot, indirect immunofluorescence, and enzyme-linked immunosorbent assay (ELISA) analyses. ELISA was used to identify whether the four VHHs were specific for PPRV, and VHH neutralization tests were carried out. ELISA and western blot analyses were used to identify which PPRV protein was targeted by VHH2. Results: The PPRV cDNA library was constructed successfully. The library capacity was greater than 2.0 × 10⁶ cfu/mL, and the inserted fragment size was approximately 400 bp to 2000 bp. The average length of the cDNA library fragment was about 1000 bp, and the recombination rate was approximately 100%. Four single-domain antibody sequences were selected, and proteins expressed in the supernatant were obtained. The four VHHs were shown to have biological activity, close affinity to PPRV, and no cross-reaction with common sheep diseases. All four VHHs had neutralization activity, and VHH2 was specific to the PPRV M protein. Conclusions: The results of this preliminary research of PPRV VHHs showed that four screened VHH antibodies could be useful in future applications. This study provided new materials for inclusion in PPRV research.

• Genomics & Informatics
• /
• v.18 no.4
• /
• pp.42.1-42.9
• /
• 2020

Chromatin immunoprecipitation coupled with high-throughput DNA sequencing (ChIP-Seq) is a powerful technology to profile the location of proteins of interest on a whole-genome scale. To identify the enrichment location of proteins, many programs and algorithms have been proposed. However, none of the commonly used peak calling programs could accurately explain the binding features of target proteins detected by ChIP-Seq. Here, publicly available data on 12 histone modifications, including H3K4ac/me1/me2/me3, H3K9ac/me3, H3K27ac/me3, H3K36me3, H3K56ac, and H3K79me1/me2, generated from a human embryonic stem cell line (H1), were profiled with five peak callers (CisGenome, MACS1, MACS2, PeakSeq, and SISSRs). The performance of the peak calling programs was compared in terms of reproducibility between replicates, examination of enriched regions to variable sequencing depths, the specificity-to-noise signal, and sensitivity of peak prediction. There were no major differences among peak callers when analyzing point source histone modifications. The peak calling results from histone modifications with low fidelity, such as H3K4ac, H3K56ac, and H3K79me1/me2, showed low performance in all parameters, which indicates that their peak positions might not be located accurately. Our comparative results could provide a helpful guide to choose a suitable peak calling program for specific histone modifications.

• Geomechanics and Engineering
• /
• v.29 no.3
• /
• pp.311-320
• /
• 2022

TBM concrete segment requires a higher level of material properties compared to general concrete structures due to difficulties in maintenance and uncertainty in ground conditions. In this regard, recently, as one of the methods to achieve enhancement effect on concrete strength, many researchers have been focusing on adding CNTs to concrete mixture. However, even CNTs do not compensate the weakness that concrete exhibits brittle behavior after cracking. Separately, over the past few decades, a number of studies have been conducted on fiber reinforced concrete which exhibits ductile behavior due to fibers bridging cracks. However, only limited studies have been conducted to employ the advantages of the both materials together. In this study, an experimental program has been conducted to investigate the effect of CNTs on the workability and the compressive behavior of PVA-ECC which exhibits ductile tensile behavior with well-distributed cracks even without a conventional rebar. In addition to the compression test, SEM analysis has been also conducted for detailed investigation in the microstructure. The variable was the CNTs mix ratio, which were set to 0.00, 0.25, and 0.50 wt.% to the binding materials. It was observed though the test results that as the CNTs mix ratio increased, the workability considerably decreased with the reduced slump and slump flow. From the compression test results, it was also investigated that the compressive behavior was improved since the compressive strength, the strain corresponding to the compressive strength, and the modulus of elasticity increased with an increase of CNTs mix ratio. The contents of this paper will be useful for relevant research areas such as fiber reinforced concrete with CNTs which might be applied for high performance TMB concrete segments.

• Journal of Life Science
• /
• v.18 no.12
• /
• pp.1733-1738
• /
• 2008

SLC6A18, one of the neurotransmitters, was reported the possible relationship to hypertension, and it contained eight blocks of minisatellites. In this study, SLC6A18-MS5 sequence which showed the highest heterozygosity among seven minisatellites was analyzed using the Transfac software, the putative binding sites for the transcription factor Pax4 and HNF4 were discovered as a result. The HNF4 is involved in the diabetes pathway and suggested the relationship to hypertension. Thus, we investigated the putative functional significance of allelic variation in this minisatellites with respect to susceptibility for hypertension. To address this possibility, we analyzed genomic DNA from the blood of 301 hypertension-free controls and 184 cases with hypertension. A statistically significant association was not identified between the allelic distribution of SLC6A18-MS5 and occurrence of hypertension. We then examined the meiotic segregation of SLC6A18-MS5 and it was transmitted following Mendelian inheritance. Therefore, this locus could be useful markers for paternity mapping and DNA fingerprinting. Moreover, we undertook a comprehensive analysis of the genomic sequence to address the evolutionary events of these variable repeats. SLC6A18 minisatellites regions are only conserved in human and primates. This result suggestedthat intronic minisatellites analysis is powerful evolution marker for the non-coding regions in primates and can provide a great insight to the molecular evolution of repeated region in primates.