• BMB Reports
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• 제45권6호
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• pp.331-336
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• 2012

The retinoid-related orphan nuclear receptor gamma ($ROR{\gamma}$) plays critical roles in regulation of development, immunity and metabolism. As transcription factor usually forms a protein complex to function, thus capturing and dissecting of the $ROR{\gamma}$ protein complex will be helpful for exploring the mechanisms underlying those functions. After construction of the recombinant tandem affinity purification (TAP) plasmid, pMSCVpuro $ROR{\gamma}$-CTAP(SG), the nuclear localization of $ROR{\gamma}$-CTAP(SG) fusion protein was verified. Following isolation of $ROR{\gamma}$ protein complex by TAP strategy, seven candidate interacting proteins were identified. Finally, the heat shock protein 90 (HSP90) and receptor-interacting protein 140 (RIP140) were confirmed to interplay with $ROR{\gamma}$ by co-immunoprecipitation. Interference of HSP90 or/and RIP140 genes resulted in dramatically decreased expression of CYP2C8 gene, the $ROR{\gamma}$ target gene. Data from this study demonstrate that HSP90 and RIP140 proteins interact with $ROR{\gamma}$ protein in a complex format and function as co-activators in the $ROR{\gamma}$-mediated regulatory processes of HepG2 cells.

• Animal cells and systems
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• 제2권4호
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• pp.539-543
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• 1998

Hrp1, of Schizosaccharomyces pombe, is a new member of the SW12/SNF2 protein family that contains a chromodomain and a DNA binding domain as well as ATPase/7 helicase domains. This configuration suggests that Hrp1 could be a homolog of mouse CHD1, which is thought to function in altering the chromatin structure to facilitate gene expression. To understand the enzymatic nature of Hrp1 we purified the 6-Histidine-tagged Hrp1 protein (6$\times$His-Hrp1) to homogeneity from a S. pombe Hrp1-overexpressing strain and hen examined its biochemical properties. We demonstrate that the purified 6$\times$His-Hrp1 protein exhibited a DNA-binding activity with a moderate preference to the (A＋T)-rich tract in double-stranded NA via a minor groove interaction. However, we failed to detect any intrinsic DNA helicase activity from the purified Hrp1 like other SW12/SNF2 proteins. These observations suggest that the DNA binding activities of Hrp1 may be involved in the remodeling of the chromatin structure with DNA-dependent ATPase. We propose that Hrp1 may function in heterochromatins as other proteins with a chromo- or ATPase/helicase domain and play an important role in the determination of chromatin architecture.

• Journal of Plant Biotechnology
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• 제37권2호
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• pp.196-204
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• 2010

With the completion of genome sequencing of several organisms, attention has been focused to determine the function and functional network of proteins by proteome analysis. The recent techniques of proteomics have been advanced quickly so that the high-throughput and systematic analyses of cellular proteins are enabled in combination with bioinformatics tools. Furthermore, the development of proteomic techniques helps to elucidate the functions of proteins under stress or diseased condition, resulting in the discovery of biomarkers responsible for the biological stimuli. Ultimate goal of proteomics orients toward the entire proteome of life, subcellular localization, biochemical activities, and their regulation. Comprehensive analysis strategies of proteomics can be classified as three categories: (i) protein separation by 2-dimensional gel electrophoresis (2-DE) or liquid chromatography (LC), (ii) protein identification by either Edman sequencing or mass spectrometry (MS), and (iii) quanitation of proteome. Currently MS-based proteomics turns shiftly from qualitative proteome analysis by 2-DE or 2D-LC coupled with off-line matrix assisted laser desorption ionization (MALDI) and on-line electrospray ionization (ESI) MS, respectively, to quantitative proteome analysis. Some new techniques which include top-down mass spectrometry and tandem affinity purification have emerged. The in vitro quantitative proteomic techniques include differential gel electrophoresis with fluorescence dyes, protein-labeling tagging with isotope-coded affinity tag, and peptide-labeling tagging with isobaric tags for relative and absolute quantitation. In addition, stable isotope labeled amino acid can be in vivo labeled into live culture cells through metabolic incorporation. MS-based proteomics extends to detect the phosphopeptide mapping of biologically crucial protein known as one of post-translational modification. These complementary proteomic techniques contribute to not only the understanding of basic biological function but also the application to the applied sciences for industry.

• Genomics & Informatics
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• 제13권2호
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• pp.53-59
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• 2015

In developing countries threat of cholera is a significant health concern whenever water purification and sewage disposal systems are inadequate. Vibrio cholerae is one of the responsible bacteria involved in cholera disease. The complete genome sequence of V. cholerae deciphers the presence of various genes and hypothetical proteins whose function are not yet understood. Hence analyzing and annotating the structure and function of hypothetical proteins is important for understanding the V. cholerae. V. cholerae O139 is the most common and pathogenic bacterial strain among various V. cholerae strains. In this study sequence of six hypothetical proteins of V. cholerae O139 has been annotated from NCBI. Various computational tools and databases have been used to determine domain family, protein-protein interaction, solubility of protein, ligand binding sites etc. The three dimensional structure of two proteins were modeled and their ligand binding sites were identified. We have found domains and families of only one protein. The analysis revealed that these proteins might have antibiotic resistance activity, DNA breaking-rejoining activity, integrase enzyme activity, restriction endonuclease, etc. Structural prediction of these proteins and detection of binding sites from this study would indicate a potential target aiding docking studies for therapeutic designing against cholera.

• 한국환경보건학회지
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• 제23권2호
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• pp.89-97
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• 1997

Most of urban streams in Korea have been changed channel forms and suffered from direct inflow of domestic sewage, etc. Therefore, maintenance of structure and function of those ecosystem are hard. The present study was carried out to examine the life survival maintenance ability of the stream by community analysis of aquatic animals in typical urban stream (Jungrang stream) in Seoul. The aquatic animals were composed of 31 species, 18 families, 8 orders, 5 classes in 4 phyla. Seasonal species number showed big fluctuation between 8 species in Winter and 24 species in Autumn. Major dominant species in Jungrang stream were Tubificidae sp.1, Chironomidae sp.1, Chironomidae sp.2 and Physa acuta, and above endurance species for water pollution occupied very high dominance indices. But, Cercion hieroglyphicum, Ischnura asiatica, Rantra chinensis, Herochares striatus, Agabus japonicus in benthic macroinvertebrates of a few individuals are appeared. Also, fry of Carassius auratus and Silurus asotus in fish are occurred. Therefore, we can be inferred on posibility of growth and spawning of above species in the stream. Jungrang stream has a small quantity of natural riffle areas, ponds and watergrass areas by channel form of water course. Aquatic animals in Jungrang stream has been suffered by reduction of self-purification reaction ability and have mass production of attached algae on the stream bed. For analysis of fluctuation of life survival maintenance ability in Jungrang stream, long-term survey is needed.

• 한국실내디자인학회논문집
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• 제20권6호
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• pp.79-87
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• 2011

Spatial perception and spatial structure that focus on psychological effects produce a real force through the medium of space that can control human actions, even their psychology. The job of understanding the characteristics and effects of architectural spaces that recognize the relationship between architecture and human beings, including the psychological dimension, is an alternative search for quality spaces that can increase the mutual relationship between space and human beings. This paper introduces two propositions called "space" and "psychology" in order to discover a meta-pattern connecting space and the human mind with the aim of systematizing that internal network and establishing a new architectural system concerning space and human beings. This paper also proposes a method of accessing physical spaces that can affect psychological states through a conceptual substitution called "extension," with the aim of discovering the implications inherent in such extensive relationships and proposing a methodology of organizing psychological spaces based on the characteristics of that extensive connection. The means of extensively connecting psychological spaces were classified into the three categories of memory system, sensory system, and motor system, and their corresponding extensive connection characteristics called "simultaneous relativity," "non-mediated immediacy," and "purification process" were also derived. These characteristics accelerate the changes in psychological intensity and function as principles that organize psychological space.

• 분석과학
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• 제17권5호
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• pp.388-392
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• 2004

genomics의 정보해석이나 신경전달물질 또는 약의 전달체계에서 아주 중요한 역할을 담당하는 막단백질의 구조연구는 기존의 X-ray나 용액상 핵자기공명분광법으로 수행하기 어려우나 지방질 이분자층이나 여러분자층에서 움직이지 않게 정렬시킨 단백질시료를 이용하여 특이하게 고안된 home-built solid-state NMR probe를 이용하면 구조를 연구할 수 있다. 이 논문에서는 박테리오파지인 pf1의 growth, 분리, 정제 및 pf1에서의 coat protein의 분리, 정제과정과 최종적으로 분리 정제된 pf1의 coat protein의 인산지방질 이분자층에서의 구조를 고체상 핵자기공명 분광법을 이용하여 연구하고자 한다.

• BMB Reports
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• 제29권2호
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• pp.116-121
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• 1996

In earlier studies, the genes encoding Escherichia coli thioredoxin and Corynebacterium nephridii thioredoxin C-3 were fused via a common restriction site in the nucleotide sequence coding for the active site of the proteins to generate two chimeric thioredoxins, designated E-C3 (N to C-terminal) and C3-E. The hybrid thioredoxins were overexpressed in E. coli from the cloned chimeric thioredoxin genes by a T7 promoter/polymerase system. To investigate the structure-function relationship of thioredoxin, we purified the E-C3 hybrid thioredoxin through ammonium sulfate fractionation, DEAE-cellulose chromatography, and Sephadex G-50 gel filtration. Its purity was examined on SDS-polyacrylamide gel electrophoresis and the molecular weight of the purified E-C3 hybrid thioredoxin was estimated to be 12,000. On native polyacrylamide gels, the purified E-C3 hybrid thioredoxin shows a much lower mobility than E. coli thioredoxin. E-C3 hybrid thioredoxin exhibits a 40-fold lower catalytic efficiency with E. coli thioredoxin reductase than E. coli thioredoxin. It was shown to catalyze the reduction of insulin disulfide by dithiothreitol. The purified E-C3 hybrid thioredoxin was also characterized in other aspects.

• BMB Reports
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• 제28권2호
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• pp.129-137
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• 1995

The ryanodine-receptor $Ca^{2+}$ release channel protein in the sarcoplasmic reticulum membrane of rabbit skeletal muscle plays an important role in muscle exitation-contraction (E-C) coupling. Various types of detergents were tested, including Chaps, cholate, octylglucoside, Zwittergents, Mega-9, Lubrol PX, and Triton X-100 for solubilization of this protein. Among these, Chaps and Triton X-100 were found to optionally solubilize the channel complex. Optimum conditions for this solubilization were pH 7.4 with a salt concentration of 1 M. The addition of phospholipid in the solubilization step helped in stabilizing the protein. The purification of the receptor was performed using sucrose density gradient centrifugation. Various methods [dilution, freeze-thaw, adsorption (Biobeads), and dialysis] were investigated to incorporate the Chaps-solubilized and purified $Ca^{2+}$ release channel protein into liposomes made from different types of phospholipids. Of these, a combined method consisting of a dialysis, freeze-thaw and sonication steps yielded the best results. Reconstituted vesicles produced by this method with 95% phosphatidylcholine (from soybean extract) had good function.

• 한국수산과학회지
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• 제20권2호
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• pp.136-145
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• 1987

말똥 성게를 인공 수정시킨 후 column chromatography법으로 DNA polymerase $\alpha$를 분리 정제하였다. Sephadex G-200과 SDS polyacryamise gel electophoresis에 의한 DNA Polymerase $\alpha$의 분자량은 약 $137,000\~138,000$이였다. 효소활성의 최적 pH는 7.4였고, 칼슘이온 20mM, 나트륨이온 25mM에서 활성이 높았고, 마그네슘 이온은 10 mM 일 때 활성이 높았다. DNA polymerase $\alpha$는 N-ethylmaleimide, aphidicolin, cytosin $\beta-D-arabinofuranoside$ 5'-triphosphate (ara CTP)와 phosphonoacetic acid체 의하여 활성이크게 저하되었다.