• Biotechnology and Bioprocess Engineering:BBE
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• v.1 no.1
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• pp.9-12
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• 1996

In order to produce the single chain precursor of a novel human insulin analogue, (B30-Homoserine) insulin, the fermentative behaviors of Escherichia coli JM103 were studied, which harbors pKBA plasmid carrying a hybrid gene in which the gene for a single chain precursor was fused with lacZ gene under tac promoter. The maximal induction of gene expression was achieved when more than 0.05 mM of isopropyl-$\beta$-D-thiogalactopyranoside(IPTG) was supplemented to fermentation medium after 4 h cultivation of E. coli, and followed by longer than 2-h fermentation. The hybrid protein of the single chain insulin precursor was isolated from cytoplasmic inclusion bodies by dissolving in 8M urea solution, and purified through DEAE-Sephacel and Sephadex G-200 column chromatographies with a recovery of 35%. The finally purified hybrid protein showed a single band on sodium dodecyl sulfate-polyacrylamide gel.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.660-664
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• 2005

We have previously isolated specific RNA aptamers with high affinity against the helicase domain of hepatitis C virus (HCV) nonstructural protein 3 (NS3). The RNA aptamers competitively and efficiently inhibited the helicase activity, partially impeding HCV replicon replication in human hepatocarcinoma cells. In this study, the RNA aptamers were tested for binding to the HCV NS3 proteins in eukaryotic cells, using a yeast three-hybrid system. The aptamers were then recognized by the HCV NS3 proteins when expressed in the cells, while the antisense sequences of the aptamers were not. These results suggest that the in vitro selected RNA aptamers can also specifically bind to the target proteins in vivo. Consequently, they could be potentially utilized as anti-HCV lead compounds.

• Proceedings of the Botanical Society of Korea Conference
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• 1985.08b
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• pp.73-81
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• 1985

We have previously isolated OsMADS4 gene that is a member of the class B MADS box genes from rice. In this study, another member of the class B MADS box genes was isolated from rice flower by the yeast two-hybrid screening method using OsMADS4 as bait. RNA blot analyses revealed that the clone, OsMADS16, was expressed in the second and third whorls, whereas the OsMADS4 transcripts were present in the second, third, and fourth whorls. These expression patterns of the OsMADS16 and OsMADS4 genes are very similar with those of AP3 and PI, the class B genes of Arabidopsis, respectively. In the yeast two-hybrid system, OsMADS4 interacted only with OsMADS16 among several rice MADS genes investigated, suggesting that OsMADS4 and OsMADS16 function as a heterodimer in specifying sepal and petal identities. We have also isolated OsMADS6 gene using OsMADS1 as a probe. Both are members of the AGL2 MADS family. Various MADS genes that encode for protein-protein interaction partners of the OsMADS6 protein were isolated by the yeast two-hybrid screening method. A majority of these genes belong to the AGL2 family. Sequence Homology, expression pattern, and ectopic expression phenotypes indicated that one of the interaction partners, OsMADS14, appears to be homologous to API, the class A MADS gene of Arabidopsis.

• Journal of Internet Computing and Services
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• v.16 no.2
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• pp.117-125
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• 2015

The information about protein structure gives the clues for the function of protein. It is needed for the improvement for the efficacy and fast development of protein drugs. So, the studies visualizing the structure of protein effectively increase. Most studies of visualization focus on the structural prediction for protein or the improvement on the rendering speed. However, studies of information delivery depending on the form of protein visualization are very limited. The major objective of this study is to analyze the information representation goodness-of-fit for the patterns of the hybrid visualization with primary and secondary structures of protein. Those hybrid visualizations included the patterns which updated current representative visualization services, Chimera, PDB and Cn3D. Information factor to analyze information representation goodness-of-fit is assorted by protein primary structure, secondary protein structure, the location of amino acid and ratio information about protein secondary structure, based on the result of subject-analysis. Subject is the group of experts who are involved in protein drug development over 5 years. The result of this study shows the meaningful difference in the information representation goodness-of-fit by the patterns of hybrid visualization and proves the difference in the information by the pattern of visualization.

• 한국약용작물학회:학술대회논문집
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• 2011.04a
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• pp.472-473
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• 2011

• BMB Reports
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• v.43 no.6
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• pp.432-437
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• 2010

LBH is a transcription factor as a candidate gene for CHD associated with partial trisomy 2p syndrome. To identify potential LBH-interacting partners, a yeast two-hybrid screen using LBH as a bait was performed with a human heart cDNA library. One of the clones identified encodes ${\alpha}B$-crystallin. Co-immunoprecipitation and GST pull-down assays showed that LBH interacts with ${\alpha}B$-crystallin, which is further confirmed by mammalian two-hybrid assays. Co-localization analysis showed that in COS-7 cells, ${\alpha}B$-crystallin that is cytoplasmic alone, accumulates partialy in the nucleus when co-transfected with LBH. Transient transfection assays indicated that overexpression of LBH or ${\alpha}B$-crystallin reduced the transcriptional activities of p53 and p21, respectively, Overexpression of both ${\alpha}B$-crystallin and LBH together resulted in a stronger repression of the transcriptional activities of p21 and p53. These results showed that the interaction of LBH and ${\alpha}B$-crystallin may inhibit synergistically the transcriptional regulation of p53 and p21.

• 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.

• Biomolecules & Therapeutics
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• v.4 no.4
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• pp.398-401
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• 1996

The glucose transporters found in the plasma membrane of all animal cells are known to have 12 putative transmembrane domains. Among 7 cytoplasmic loops, the fourth loop is the largest one. Since previous studies showed that cofilin, an actin-modulating protein, was found to interact with the largest cytoplasmic loop of (Na, K)ATPase, we tested if cofilin interacts with the largest cytoplasmic loop of Glut4. We demonstrated by the two-hybrid system that the largest cytoplasmic loop of Glut4 did not show any interaction with cofilin, suggesting that cofilin is not required for the membrane targeting process of other membrane proteins but only for a P-type ATPase.

• Animal cells and systems
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• v.6 no.2
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• pp.181-181
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• 2002

Nek2 is a mammalian protein kinase that is structurally homologous to NIMA, a mitotic regulator in Aspergillus nidulans. We recently observed that the Nek2 protein was localized in multiple sites within a cell in a cell cycle state-specific manner. This suggests that Ndk2 is involved in diverse cellular functions during the cell cycle progression. To have a better understanding on cellular functions in which Nek2 participates, we carried out yeast two-hybrid screening and isolated six candidate clones whose products interact with Nek2. Most of Nek2-interacting proteins (NIPs) appear cytoplasmic, suggesting that Nek2 is involved in cellular functions in cytoplasm. Further experiments are under progress to confirm their interactions with Nek2 and to understand their biological significance.

• BMB Reports
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• v.38 no.4
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• pp.440-446
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• 2005

A cDNA that was rapidly induced upon abscisic acid, cold, drought, mechanical wounding and to a lesser extent, by high salinity treatment, was isolated from Arabidopsis seedlings. It was classified as DREB subfamily member based on multiple sequence alignment and phylogenetic characterization. Since it encoded a protein with a typical ERF/AP2 DNA-binding domain and was closely related to the TINY gene, we named it TINY2. Gel retardation assay revealed that TINY2 was able to form a specific complex with the previously characterized DRE element while showed only residual affinity to the GCC box. When fused to the GAL4 DNA-binding domain, either full-length or its C-terminus functioned effectively as a trans-activator in the yeast one-hybrid assay while its N-terminus was completely inactive. Our data indicate that TINY2 could be a new member of the AP2/EREBP transcription factor family involved in activation of down-stream genes in response to environmental stress.