• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.4
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• pp.263-267
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• 2003

Magnetic nanoparticles prepared from an alkaline solution of divalent and trivalent iron ions could covalently bind protein via the activation of Nethyl-N-(3-dimethylaminopropyl) carbodiimide (EDC). Trypsin and avidin were taken as the model proteins for the formation of protein-nanoparticle conjugates. The immobilized yield of protein increased with molar ratio of EDC/nanoparticie. Higher concentrations of added protein could yield higher immobilized protein densities on the particles. In contrast to EDC, the yields of protein immobilization via the a ctivation of cyanamide were relatively lower. Nanoparticles bound with avidin could attach a single-stranded DNA through the avidin-biotin interaction and hybridize with a DNA probe. The DNA hybridization was confirmed by fluorescence microscopy observations. Immobilized DNA on nanoparticles by this technique may have widespread applicability to the detection of specific nucleic acid sequence and targeting of DNA to particular cells.

• Archives of Pharmacal Research
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• v.9 no.3
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• pp.145-152
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• 1986

Magnetically reponsive gelatin microspheres for the targeting of drugs have been prepared using a water-in-oil emulsion technique with chemical cross-linking of the protein. The manufacturing variables affecting microsphere size, size distribution and surface characteristics have been examined as well as the magnetic responsiveness in vitro. Sesame oil was utilized for non-aqueous phase and magentic gelatin microspheres of different size from 1. 89 to 14.88 $\mu\textrm{m}$ in mean diameter could be obtained with variation of HLB values of non-ionic surfactants. The content of magnetite which uniformly distributed throughout the microspheres was 26.7% (w/w). It was possible to control the localization of magnetic gelatin microspheres at specific sites within capilary models by using external magnetic field of under 5K gauss.

• Korean Journal of Plant Tissue Culture
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• v.27 no.4
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• pp.283-297
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• 2000

Seeds are an ideal repository for recombinant proteins in molecular farming applications. However, in order to use plant seeds efficiently for the production of such proteins, it is necessary to understand a number of fundamental biological properties of seeds. This includes a full understanding of promoters which function in a seed-specific manner, the subcellular targeting of the desired polypeptide and the final form in which a protein is stored. Once a biologically active protein has been deposited in a seed, it is also critical that the protein can be extracted and purified efficiently. In this review, these issues are examined critically to provide a number of approaches which may be adopted for production of recombinant proteins in plants. Particular attention is paid to the relationship between subcellular localization and protein extraction and purification. The robustness and flexibility of seed-based production is illustrated by examples close to or already in commercial production.

• BMB Reports
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• v.31 no.2
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• pp.196-200
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• 1998

Zeins, maize storage proteins, are retained in the endoplasmic reticulum (ER) during the subcellular targeting process without the ER retention signal. Circumstantial data indicate that the 27K zein is an ER transmembrane protein. The potential transmembrane domain may permit the 27K zein to remain in the ER. This study investigated the potential transmembrane feature by employing alkaline extraction, proteinase K digestion, and surface biotinylation on isolated intact protein bodies. These assays consistently support the possibility of the 27K zein as a transmembrane protein. The 27K zein polypeptide was shown to be associated with alkali-stripped membranes. The polypeptide was digested by proteinase K to a smaller fragment. According to surface biotinylation, the 27K zeins was labeled to the exclusion of other classes of zeins. This study, therefore, concludes that the 27K zein has an ER transmembrane domain, which may serve as an anchor for zeins' ER retention.

• BMB Reports
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• v.54 no.7
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• pp.380-385
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• 2021

Proper targeting of the βPAK-interacting exchange factor (βPIX)/G protein-coupled receptor kinase-interacting target protein (GIT) complex into distinct cellular compartments is essential for its diverse functions including neurite extension and synaptogenesis. However, the mechanism for translocation of this complex is still unknown. In the present study, we reported that the conventional kinesin, called kinesin-1, can transport the βPIX/GIT complex. Additionally, βPIX bind to KIF5A, a neuronal isoform of kinesin-1 heavy chain, but not KIF1 and KIF3. Mapping analysis revealed that the tail of KIF5s and LZ domain of βPIX were the respective binding domains. Silencing KIF5A or the expression of a variety of mutant forms of KIF5A inhibited βPIX targeting the neurite tips in PC12 cells. Furthermore, truncated mutants of βPIX without LZ domain did not interact with KIF5A, and were unable to target the neurite tips in PC12 cells. These results defined kinesin-1 as a motor protein of βPIX, and may provide new insights into βPIX/GIT complex-dependent neuronal pathophysiology.

• Genomics & Informatics
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• v.20 no.3
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• pp.33.1-33.17
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• 2022

Nervous necrosis virus (NNV) is a deadly infectious disease that affects several fish species. It has been found that the NNV utilizes grouper heat shock cognate protein 70 (GHSC70) to enter the host cell. Thus, blocking the virus entry by targeting the responsible protein can protect the fishes from disease. The main objective of the study was to evaluate the inhibitory potentiality of 70 compounds of Azadirachta indica (Neem plant) which has been reported to show potential antiviral activity against various pathogens, but activity against the NNV has not yet been reported. The binding affinity of 70 compounds was calculated against the GHSC70 with the docking and molecular dynamics (MD) simulation approaches. Both the docking and MD methods predict 4 (PubChem CID: 14492795, 10134, 5280863, and 11119228) inhibitory compounds that bind strongly with the GHSC70 protein with a binding affinity of -9.7, -9.5, -9.1, and -9.0 kcal/mol, respectively. Also, the ADMET (absorption, distribution, metabolism, excretion, and toxicity) properties of the compounds confirmed the drug-likeness properties. As a result of the investigation, it may be inferred that Neem plant compounds may act as significant inhibitors of viral entry into the host cell. More in-vitro testing is needed to establish their effectiveness.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.86-86
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• 2003

Research has been in progress for more than a decade to production of useful proteins by genetic modification in cattle. However, the levels of protein production in transgenic cattle have been reported very low. To enhance protein production in transgenic animal, we tried homologous recombination to donor cells for production of transgenic clone cattle through nuclear transfer procedure. Thus, we constructed the two targeting vectors of human thrombopoietin (TPO) at bovine $\beta$-casein locus using homologous recombination with 13.6 kb and 9.6 kb homology. In two targeting vectors, positive selection was through the neomycin resistance gene and negative selection was by the diphtheria toxin (DT). Gene targeting was attempted in bovine embryonic fibroblasts (bEF) and bovine ear skin fibroblasts (bESF). To determine the most appropriate concentration of neomycin for bEF and bESF, G4l8 resistance was confirmed by culturing the cells in various concentrations of the drug and both of the cells were optimally selected at $900 \mu g/ml$ of neomycin. The transfected bEF and bESF by the targeting vectors were colonized efficiently at the ratio of DNA to transfection reagent such as $4 \mu g$:2 ${mu}ell$ and $1 \mu g$:$2 \mu l$. Comparing number of healthy clones from passage 4 to passage 8, bESF (17%) persist in culture for much longer than bEF (6%). The two gene-targeted bESF clones of 30 random-integrated clones with 9.6 kb homology length were confirmed, however, nothing was out of 72 random integration clones with 13.6 kb homology length, The DT also worked more efficiently in clones transfected with the vector of 9.6 kb homology length. Our data suggests that the choice of donor cell for long culture period should be considered to obtain targeted cell clone, and the gene-targeting frequency and the DT working efficiency are dependent on the length of target homology.

• Bulletin of the Korean Chemical Society
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• v.28 no.11
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• pp.1917-1918
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• 2007

• KSBB Journal
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• v.25 no.3
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• pp.215-222
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• 2010

Monogenic diseases are resulted from modifications in a single gene of human cells. Because their treatment with pharmacological medicine have a temporary effect, continuous nursing care and retreatment are required. Gene therapy, gene targeting and induced pluripotent stem cell (iPSC) are considered permanent treatment methods of them. In gene therapy, however, retroviral vectors that have potential toxicity caused by random insertion of harmful virus are used as vehicles for transferring genetic materials. On the other hand, gene targeting could replace and remove the modified gene though homologous recombination (HR) induced by site-specific endonucleases. This short review provides a brief overview on the recently tailored endonucleses with high selectivity for HR.

• International Journal of Industrial Entomology and Biomaterials
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• v.9 no.1
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• pp.149-153
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• 2004

Here we report the molecular cloning of a LIM protein cDNA of the CRP (cysteine-rich protein) family from the mulberry longicorn beetle, Apriona, geramri. The A. germari LIM protein cDNA contains an open reading frame of 276 bp encoding 92 amino acid residues with a calculated molecular weight of approximately 10 kDa. The A. germari LIM protein contains the cysteine-rich consensus sequence of LIM domain and the glycine-rich consensus sequence observed in cysteine-rich protein family 1 (CRP1). The potential nuclear targeting signal is retained. The deduced amino acid sequence of the A. germari LIM protein cDNA showed 81 % identity to both Bombyx mori muscle LIM protein (Mlp) and Drosophila melanogaster Mlp60A and 77% to Epiblema scudderiana Mlp. Northern blot analysis showed that A. germari LIM protein is highly expressed in epidermis and muscle, and less strongly in midgut, but not in the fat body.