• BMB Reports
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• v.47 no.2
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• pp.92-97
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• 2014

We have examined the effect of bithorax complex genes on the expression of castor gene. During the embryonic stages 12-15, both Ultrabithorax and abdominal-A regulated the castor gene expression negatively, whereas Abdominal-B showed a positive correlation with the castor gene expression according to real-time PCR. To investigate whether ABD-B protein directly interacts with the castor gene, electrophoretic mobility shift assays were performed using the recombinant ABD-B homeodomain and oligonucleotides, which are located within the region 10 kb upstream of the castor gene. The results show that ABD-B protein directly binds to the castor gene specifically. ABD-B binds more strongly to oligonucleotides containing two 5'-TTAT-3' canonical core motifs than the probe containing the 5'-TTAC-3' motif. In addition, the sequences flanking the core motif are also involved in the protein-DNA interaction. The results demonstrate the importance of HD for direct binding to target sequences to regulate the expression level of the target genes.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.6
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• pp.333-339
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• 2005

B/K protein is a novel protein containing double C2-like domains. We examined the specific signaling pathway that regulates the transcription of B/K in PC12 cells. When the cells were treated with forskolin ($50{\mu}M$), B/K mRNA and protein levels were time-dependently decreased, reaching the lowest level at 3 or 4 hr, and thereafter returning to the control level. Chemicals such as dibutyryl-cAMP, cellpermeable cyclic AMP (cAMP) analogue and CGS21680, adenosine receptor $A_{2A}$ agonist, also repressed the B/K transcription. However, 1,9-dideoxyforskolin did not show inhibitory effect on B/K transcription, suggesting direct involvement of cAMP in the forskolin-induced inhibition of B/K transcription. Effect of forskolin, dibutyryl cAMP and CGS21680 was significantly reduced in PKA-deficient PC12 cell line (PC12-123.7). One cAMP-response element (CRE)-like sequence (B/K CLS) was found in the promoter region of B/K DNA, and electrophoretic mobility shift assay indicated its binding to CREM and CREB. Forskolin significantly suppressed the promoter activity in CHO-K1 cells transfected with the constructs containing B/K CLS, but not with the construct in which B/K CLS was mutated (AC：TG). Taken together, we suggest that the transcription of B/K gene in PC12 cells may be regulated by PKA-dependent mechanism.

• Toxicological Research
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• v.33 no.3
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• pp.211-218
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• 2017

Cytochrome P450 1B1 (CYP1B1) acts as a hydroxylase for estrogen and activates potential carcinogens. Moreover, its expression in tumor tissues is much higher than that in normal tissues. Despite this association between CYP1B1 and cancer, the detailed molecular mechanism of CYP1B1 on cancer progression in HeLa cells remains unknown. Previous reports indicated that the mRNA expression level of Herc5, an E3 ligase for ISGylation, is promoted by CYP1B1 suppression using specific small interfering RNA, and that ISGylation may be involved in ubiquitination related to ${\beta}-catenin$ degradation. With this background, we investigated the relationships among CYP1B1, Herc5, and ${\beta}-catenin$. RT-PCR and western blot analyses showed that CYP1B1 overexpression induced and CYP1B1 inhibition reduced, respectively, the expression of $Wnt/{\beta}-catenin$ signaling target genes including ${\beta}-catenin$ and cyclin D1. Moreover, HeLa cells were treated with the CYP1B1 inducer $7,12-dimethylbenz[{\alpha}]anthracene$ (DMBA) or the CYP1B1 specific inhibitor, tetramethoxystilbene (TMS) and consequently DMBA increased and TMS decreased ${\beta}-catenin$ and cyclin D1 expression, respectively. To determine the correlation between CYP1B1 expression and ISGylation, the expression of ISG15, a ubiquitin-like protein, was detected following CYP1B1 regulation, which revealed that CYP1B1 may inhibit ISGylation through suppression of ISG15 expression. In addition, the mRNA and protein expression levels of Herc5 were strongly suppressed by CYP1B1. Finally, an immunoprecipitation assay revealed a direct physical interaction between Herc5 and ${\beta}-catenin$ in HeLa cells. In conclusion, these data suggest that CYP1B1 may activate $Wnt/{\beta}-catenin$ signaling through stabilization of ${\beta}-catenin$ protein from Herc5-mediated ISGylation for proteosomal degradation.

• The KIPS Transactions:PartB
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• v.13B no.1 s.104
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• pp.1-6
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• 2006

The prediction of protein function basically make use of a protein-protein interaction map based on the concept of guilt-by-association. The method however cannot determine the functions of proteins in case that the target protein does not interact with proteins with known functions directly. This paper studies protein function prediction considering the given problem as a K-class classification problem and proposes a predictive approach utilizing a modular neural network. The proposed method uses interaction data and protein related attributes as well. The experimental results demonstrate that the proposed approach can predict the functional roles of Yeast proteins whose interaction knowledge is not known and shows better performance than the graph-based models that use protein interaction data.

• The Korean Journal of Mycology
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• v.16 no.3
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• pp.121-127
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• 1988

For selective purification of protein in Pleurotus cornucopiae (Per.) Rolland, affinity chromatography was performed by benzoyl-AH-Sepharose 4B gel synthesized using AH-Sepharose 4B with starting materials. Ligand capacity of benzoyl group was 9.3 micromole per milliliter of gel. Total apparent molecular weight of affinity protein was 255KD, which were protein complex of 29.5, 31.5 34.0, 71.0 and 89.0KD, respectively. The contents of nonpolar, polar, positively charged, and negatively charged amino acid were 45.68%, 26.93%, 11.81% and 15.58%, respectively. The nonpolar protein was selectively purified by hydrophobic ligand of benzoyl group of gel.

• Journal of Life Science
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• v.23 no.8
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• pp.978-983
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• 2013

Kinesin-II, a molecular motor, consists of two different motor subunits, KIF3A and KIF3B, and one large kinesin superfamily-associated protein 3 (KAP3), forming a heterotrimeric complex. KAP3 is associated with the tail domains of motor subunits. However, its exact role remains unclear. Here, we demonstrated KAP3 binding to the carboxyl (C)-terminal tail region of HS-associated protein X-1 (HAX-1). HAX-1 bound to the C-terminal region of KAP3, but not to KIFs (KIF3A, KIF3B, and KIF5B) and the kinesin light chain (KLC) in the yeast two-hybrid assays. The interaction was further confirmed in the glutathione S-transferase (GST) pull-down assay and by co-immunoprecipitation. Anti- HAX-1 antibody as well as anti-KIF3A antibody co-immunoprecipitated KIF3B and KAP3 from mouse brain extracts. These results suggest that KAP3 could mediate the interaction between Kinesin-II and HAX-1.

• Journal of Embryo Transfer
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• v.28 no.1
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• pp.57-61
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• 2013

Microtubule-associated protein 1B (MAP1B), a member of MAP1 family, plays a key role in neuronal development. MAP1B binds to many kinds of proteins directly or indirectly. This study was performed to investigate whether MAP1B interacts with GAPDH in bovine follicles using immunoprecipitation (IP) with Western blot analysis and immunohistochemisty. The mRNA expressions of MAP1B and glyceraldehydes 3-phosphate dehydrogenase (GAPDH) were down-regulated in bovine follicular cystic follicles (FCF). In parallel with the mRNA levels, their protein levels were also down-regulated in FCFs. In addition, MAP1B and GAPDH were co-localized at the cytoplasm of follicles. IP with Western blot analysis showed that MAP1B bound to GAPDH in normal follicles, but their binding was absent in FCFs, suggesting a low level of MAP1B and/or GAPDH expressions in FCFs. Taken together, these results suggest that MAP1B interacted with GAPDH may play a role in bovine follicle development, and that GAPDH does not function always as a loading control in bovine follicles.

• Bulletin of the Korean Chemical Society
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• v.29 no.8
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• pp.1479-1484
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• 2008

Protein tyrosine phosphatase (PTP) 1B is the superfamily of PTPs and a negative regulator of multiple receptor tyrosine kinases (RTKs). Inhibition of protein tyrosine phosphatase 1B (PTP1B) has been proposed as a strategy for the treatment of type 2 diabetes and obesity. Recently, it has been reported that amentoflavone, a biflavonoid extracted from Selaginella tamariscina, inhibited PTP1B. In the present study, docking model between amentoflavone and PTP1B was determined using automated docking study. Based on this docking model and the interactions between the known inhibitors and PTP1B, we determined multiple pharmacophore maps which consisted of five features, two hydrogen bonding acceptors, two hydrogen bonding donors, and one lipophilic. Using receptor-oriented pharmacophore-based in silico screening, we searched the biflavonoid database including 40 naturally occurring biflavonoids. From these results, it can be proposed that two biflavonoids, sumaflavone and tetrahydroamentoflavone can be potent allosteric inhibitors, and the linkage at 5',8''-position of two flavones and a hydroxyl group at 4'-position are the critical factors for their allosteric inhibition. This study will be helpful to understand the mechanism of allosteric inhibition of PTP1B by biflavonoids and give insights to develop potent inhibitors of PTP1B.

• Journal of Dairy Science and Biotechnology
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• v.35 no.4
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• pp.262-265
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• 2017

Bombyx mori (silkworm pupae) is a unique and biologically significant insect, which is a recognized source of high quality protein that provides all the essential amino acids required for human health. Recently, many studies have focused on various biomedical applications of B. mori proteins. The purpose of this study was to manufacture protein-fortified kefir containing different concentrations of B. mori powder according to pH and sensory evaluations. The value of the protein-fortified kefir increased but the pH decreased with increasing incubation time, indicating that the amount of B. mori powder did not affect and pH. Addition of B. mori powder also did not affect the sensory properties of overall acceptability, texture, and color compared to control group without addition of B. mori powder. However, flavor and taste were affected by increasing the amounts of B. mori powder, with a significant difference in both flavor and taste between the control and treated groups (both p<0.05). There was no significant difference in overall acceptability, texture, and color. Further studies are needed for producing kefir as a dietary supplement utilizing the functional properties of B. mori.

• Development and Reproduction
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• v.15 no.1
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• pp.25-30
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• 2011

Embryonic stem (ES) cells can self-renew maintaining the undifferentiated state. Self renewal requires many factors such as Oct4, Sox2, FoxD3, and Nanog. NF-${\kappa}B$ is a transcription factor involved in many biological activities. Expression and activity of NF-${\kappa}B$ increase upon differentiation of ES cells. Reportedly, Nanog protein directly binds to NF-${\kappa}B$ protein and inhibits its activity in ES cells. Here, we found a potential binding site of NF-${\kappa}B$ in the human Nanog promoter and postulated that NF-${\kappa}B$ protein may regulate expression of the Nanog gene. We used human embryonic carcinoma (EC) cells as a model system of ES cells and confirmed decrease of Nanog and increase of NF-${\kappa}B$ upon differentiation induced by retinoic acid. Although deletion analysis on the DNA fragment including NF-${\kappa}B$ binding site suggested involvement of NF-${\kappa}B$ in the negative regulation of the promoter, site-directed mutation of NF-${\kappa}B$ binding site had no effect on the Nanog promoter activity. Furthermore, no direct association of NF-${\kappa}B$ with the Nanog promoter was detected during differentiation. Therefore, we conclude that NF-${\kappa}B$ protein may not be involved in transcriptional regulation of Nanog gene expression in EC cells and possibly in ES cells.