• Proceedings of the Korean Biophysical Society Conference
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• 1997.07a
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• pp.9-10
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• 1997

Calmodulin: very large conformation change of helix uncoiling, hinge-bending and domain rotation. Calmodulin (CaM) is the principal Ca$\^$2＋/ -dependent regulator of a variety of important eukaryotic cellular processes. In many of these processes, calmodulin activates a plethora of target enzymes, and the calmodulin-binding domains in several targets have been shown to residue in a region of about 18-residue peptide segment.(omitted)

• Bulletin of the Korean Chemical Society
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• v.17 no.2
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• pp.182-188
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• 1996

Early attempts to identify plausible conformations of a linear octapeptide hormone, angiotensin-II (Asp-Arg-Val-Tyr-Ile-His-Pro-Phe), using various theoretical and experimental methods, have led to various conformational models. So far, no consensus has been made about the solution phase structure and the receptor binding structure of angiotensin-II. The ultimate goal for the conformation study of the peptide hormone is to develop a new potent drug. Therefore, we have devised a strategy for designing the pharmacophore by studying thermodynamically possible conformations of various kinds of angiotensin-II antagonists and angiotensin-II.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.175.2-175.2
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• 2003

Inhibition of the protein-modifying enzyme farnesyltransferase is considered as a major emerging strategy in cancer therapy because of the involvement of farnesylated proteins in oncogensis. We studied the structure-activity relationship of a novel class of CAAX-peptidomimetic farnesyltransferase inhibitors based on the benzophenone scaffold. FlexX docking of inhibitors confirmed reasonable fit of the molecule into the peptide binding site of farnesyltransferase. We also performed a virtual screening with LeadQuest chemical library databases to idenfity novel inhibitors of farnesyltransferase. (omitted)

• Molecules and Cells
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• v.37 no.3
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• pp.264-269
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• 2014

The molecular interaction between tumor suppressor p53 and the anti-apoptotic Bcl-2 family proteins plays an essential role in the transcription-independent apoptotic pathway of p53. In this study, we investigated the binding of p53 DNA-binding domain (p53DBD) with the anti-apoptotic Bcl-2 family proteins, Bcl-w, Mcl-1, and Bcl-2, using GST pull-down assay and NMR spectroscopy. The GST pull-down assays and NMR experiments demonstrated the direct binding of the p53DBD with Bcl-w, Mcl-1, and Bcl-2. Further, NMR chemical shift perturbation data showed that Bcl-w and Mcl-1 bind to the positively charged DNA-binding surface of p53DBD. Noticeably, the refined structural models of the complexes between p53DBD and Bcl-w, Mcl-1, and Bcl-2 showed that the binding mode of p53DBD is highly conserved among the anti-apoptotic Bcl-2 family proteins. Furthermore, the chemical shift perturbations on Bcl-w, Mcl-1, and Bcl-2 induced by p53DBD binding occurred not only at the p53DBD-binding acidic region but also at the BH3 peptide-binding pocket, which suggests an allosteric conformational change similar to that observed in Bcl-$X_L$. Taken altogether, our results revealed a structural basis for a conserved binding mechanism between p53DBD and the anti-apoptotic Bcl-2 family proteins, which shed light on to the molecular understanding of the transcription-independent apoptosis pathway of p53.

• Journal of Microbiology and Biotechnology
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• v.11 no.1
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• pp.153-159
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• 2001

In order to clone the peptide synthetase gene form Lysobacter lactamgenus IFO 14,288, the gene fragments were amplified using primers for the adenylation domain and the thionylation domain of the peptide synthetase genes in other organisms by polymerase chain reaction (PCR). The resulting 0.5-kb fragment was cloned in a pGEM-T vector, and the nucleotide sequences were determined. Six different PCR products were obtained; three were identified to be a part of L-$\alpha$-aminoadipyl-L-cysteinyl-D-valine (ACV) synthetase and three to be other peptide synthetases. Using each of the two different classes of PCR products as mixed probes, a cosmid library of L. lactamgenus chromosomal DNA constructed in a pHC79 vector was screened by an in situ hybridization procedure, and one positive clone was selected which was bound by peptide synthetase gene fragments as well as ACV synthetase gene fragments. The partial sequence analysis formt he obtained pPTS-5 cosmid showed th presence of more than two open reading frames. These were for two putative membrane transporters, which were homologous with several integral membrane proteins including the ABC transporter ATP-binding protein of E. coli (YbjZ) and the metal ion uptake protein of Bacillus subtilis (YvrN). A 45% homology was also found between the two transporter proteins at the carboxy terminus. Through a hydropathy analysis and transmembrane analysis. 4-5 transmembrane domains were found in these two proteins. When the genes were expressed in Escherichia coli, the gene products inhibited the hose cell growth, probably due to the disturbance of the membrane transport system.

• BMB Reports
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• v.32 no.5
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• pp.461-467
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• 1999

Recombinant Mycobacterium smegmatis expressing ovalbumin was used to immunize C57BL/6(H-$2^b$) mice, and the humoral immunity against recombinant ovalbumin was analyzed. Antibodies were purified by denatured ovalbumin-conjugated affinity chromatography. The epitopes of the antibodies were screened with a random peptide library displayed on the tip of fUSE5 filamentous phage pIII minor coat proteins. Two peptides, IRLADR and SPGAEV, were selected predominantly by the recognition of purified antibodies using biopanning methods. The composition of the peptide sequence with the primary structure of OVA revealed that the peptide sequence analogizes to INEAGR, part of the $^{323}ISQAVHAAHAEINEAGR^{339}$ sequence previously reported as the antigenic determinant for murine Band also Th cell epitopes (I-$A^d$ binding). Also, the structures of these mimotopes obtained from restrained molecular dynamic computations resulted in the formation of a $\beta$-turn proven to be a secondary structure of the parent peptide within the ovalbumin molecule, enabling us to confirm the structural similarity. This study demonstrates that immunization with recombinant M. smegmatis can generate neutralizing antibodies identical with those induced by the administration of natural antigenic proteins and supports the potential use of mycobacteria as vaccine delivery vehicles.

• Journal of Microbiology and Biotechnology
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• v.27 no.11
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• pp.1999-2009
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• 2017

The secretion efficiency of a protein in a Sec-type secretion system is mainly determined by an N-terminal signal peptide and its combination with its cognate protein. Five signal peptides, namely, two synthetic Sec-type and three Bacillus licheniformis alpha-amylase-derived signal peptides, were compared for periplasmic expression of the human growth hormone (hGH) in E. coli. Based on in silico predictions on the signal peptides' cleavage efficiencies and their corresponding mRNA secondary structures, a number of amino acid substitutions and silent mutations were considered in the modified signal sequences. The two synthetic signal peptides, specifically designed for hGH secretion in E. coli, differ in their N-terminal positively charged residues and hydrophobic region lengths. According to the mRNA secondary structure predictions, combinations of the protein and each of the five signal sequences could lead to different outcomes, especially when accessibility of the initiator ATG and ribosome binding sites were considered. In the experimental stage, the two synthetic signal peptides displayed complete processing and resulted in efficient secretion of the mature hGH in periplasmic regions, as was demonstrated by protein analysis. The three alpha-amylase-derived signal peptides, however, were processed partially from their precursors. Therefore, to achieve efficient secretion of a protein in a heterologous system, designing a specific signal peptide by using a combined approach of optimizations of the mRNA secondary structure and the signal peptide H-domain and cleavage site is recommended.

• Biomolecules & Therapeutics
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• v.19 no.1
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• pp.45-51
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• 2011

These experiments were designed to use typical makers from behaviors and molecular basis in addicted animals of morphine and cocaine. Morphine has been widely abused with a high physical dependence liability. Morphine withdrawal activates the intracellular cAMP signaling pathway and further leads to changes in the expression of the cAMP response element binding protein (CREB), which may be important to the development and expression of morphine dependence. From these experiments, repeated morphine (10 mg/kg, twice per day for 7 days) developed physical dependence. Withdrawal signs were precipitated by naloxone and also increased the expression of the CREB. In addition, repeated exposure of cocaine (15 mg/kg) to mice develops locomotor sensitization and produced lasting behavioral sensitivity. Cocaine- and amphetamine-regulated transcript peptide (CART) peptide was up-regulated by repeated administration of cocaine in the striatum. Therefore, repeated morphine induced the development of physical dependence and increased pCREB. In addition, repeated cocaine induced locomotor sensitization and over-expressed CART peptide. In conclusion, the development of physical dependence and pCREB for morphine, and locomotor sensitization and CART peptide over-expression for cocaine would be useful markers to predict the abuse potential of opioid analgesics and pychostimulant drugs in animals, respectively.

• Animal cells and systems
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• v.8 no.1
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• pp.49-55
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• 2004

In vertebrates, multisubunit cofactors regulate gene expression through interacting with cell-type- and gene-specific DNA-binding proteins in a chromatin-selective manner. ADD1/SREBP1c regulates fatty acid metabolism and insulin-dependent gene expression through binding to SRE and E-box motif with dual DNA binding specificity. Although its transcriptional and post-translational regulation has been extensively studied, its regulation by interacting proteins is not well understood. To identify cellular proteins that associate with nuclear form of ADD1/SEBP1c, we employed the GST pull-down system with Hela cell nuclei extract. In this study, we demonstrated that Ku proteins interact specifically with ADD1/SREP1c protein. GST pull-down combined with peptide sequencing analysis revealed that Ku80 binds to ADD1/SREBP1c in vitro. Additionally, western blot analysis showed that Ku70, a heterodimerizing partner of Ku80, also associates with ADD1/SREBP1c. Furthermore, co-transfection of Ku70/Ku80 with ADD1/SREBP1c enhanced the transcriptional activity of ADD1/SREBP1c. Taken together, these results suggest that the Ku proteins might be involved in the lipogenic and/or adipogenic gene expression through interacting with ADD1/SREBP1c.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.537-543
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• 2005

Subtilisin (EC 3.4.21.14) is the major extracellular alkaline serine protease of Bacillus species. Previously, we found that subtilisins did not migrate in the electrophoretic field in the Laemmili buffer system due to their high pI values (over 8.8); however, it formed a 'binding mode' at the top of the separating gel [5]. Utilizing this characteristic, four subtilisins from Bacillus sp. strains (e.g., B. subtilis 168, B. subtilis KCTC 1021, B. amyloliquefaciens KCTC 3002, and Bacillus sp. DJ-1 and DJ-4) were easily and quickly identified by an over-running electrophoretic technique with a miniscale culture supernatant (less than 20 ml) without any column chromatographic steps. Two subtilisins (DJ-l and a recombinant version) from Bacillus sp. DJ-l were characterized, and the enzymatic properties were determined by SDS-fibrin zymography and densitometric analysis. Based on this observation, the recombinant pro-subtilisin DJ-l showed the same 'binding mode,' similar to native subtilisin DJ-l. On the other hand, mature subtilisin DJ -1 without pro-peptide showed no enzymatic activity.