• BMB Reports
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• 제34권2호
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• pp.156-165
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• 2001

Leptin is a circulating non-glycosylated protein that is mainly produced in adipocytes. Leptin acts in the brain to regulate food intake and energy expenditure. Previously we reported our success in the isolation of a partial cDNA of the long form of the leptin receptor, OB-Rb, from rat spleen, and showed that leptin might also play a role in peripheral immune organs. In the present study, for the first time, the complete coding region of OB-Rb cDNA was cloned from rat splenocytes, and its nucleotide sequence was determined. The cDNA was then further expressed in E. coli and mammalian cells, thereby confirming the functional integrity of this receptor. Prokaryotically overexpressed OB-R protein was then used as an immunizing antigen in BALE/c mice to produce leptin receptor-specific antibodies. By using them, we confirmed the cell surface expression of OB-Rb in transfected CHO cells. It is our belief that the reagents, as produced in this study, will be of great use in further studies of the biological role of rat leptin.

• Biomolecules & Therapeutics
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• 제25권1호
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• pp.12-25
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• 2017

G protein-coupled receptors (GPCRs) are a family of cell-surface proteins that play critical roles in regulating a variety of pathophysiological processes and thus are targeted by almost a third of currently available therapeutics. It was originally thought that GPCRs convert extracellular stimuli into intracellular signals through activating G proteins, whereas ${\beta}$-arrestins have important roles in internalization and desensitization of the receptor. Over the past decade, several novel functional aspects of ${\beta}$-arrestins in regulating GPCR signaling have been discovered. These previously unanticipated roles of ${\beta}$-arrestins to act as signal transducers and mediators of G protein-independent signaling have led to the concept of biased agonism. Biased GPCR ligands are able to engage with their target receptors in a manner that preferentially activates only G protein- or ${\beta}$-arrestin-mediated downstream signaling. This offers the potential for next generation drugs with high selectivity to therapeutically relevant GPCR signaling pathways. In this review, we provide a summary of the recent studies highlighting G protein- or ${\beta}$-arrestin-biased GPCR signaling and the effects of biased ligands on disease pathogenesis and regulation.

• Journal of Microbiology and Biotechnology
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• 제25권11호
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• pp.1856-1862
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• 2015

In order to improve the stability of endoglucanase under thermal and acidic conditions, the endoglucanase gene was fused to the N-terminus of the Saccharomyces cerevisiae pir gene, encoding the cell wall protein PIR. The fusion gene was transformed into Pichia pastoris GS115 for expression. A resulting strain with high expression and high activity was identified by examining resistance to Geneticin 418, Congo red staining, and quantitative analysis of enzyme activity. SDS-PAGE analysis revealed that the endoglucanase was successfully displayed on the yeast cell surface. The displayed endoglucanase (DEG) showed maximum activity towards sodium carboxyl methyl cellulose at approximately 275 IU/g cell dry weight. DEG exhibited greater than 60% residual activity in the pH range 2.5-8.5, higher than free endoglucanase (FEG), which had 40% residual activity at the same pH range. The highest tolerated temperature for DEG was 70℃, much higher than that of FEG, which was approximately 50℃. Moreover, DEG showed 91.1% activity at 65℃ for 120 min, while FEG only kept 77.8% residual activity over the same period. The half-life of DEG was 270 min at 65℃, compared with only 150 min for FEG. DEG could be used repeatedly at least three times. These results suggest that the DEG has broad applications as a yeast whole-cell biocatalyst, due to its novel properties of high catalytic efficiency, acid-thermal stabilities, and reusability.

• Journal of Microbiology and Biotechnology
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• 제22권2호
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• pp.199-206
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• 2012

Naturally immobilized tannase (tannin acyl hydrolase, E.C. 3.1.1.20) has many advantages, as it avoids the expensive and laborious operation of isolation, purification, and immobilization, plus it is highly stable in adverse pH and temperature. However, in the case of cell-associated enzymes, since the enzyme is associated with the biomass, separation of the pure biomass is necessary. However, tannic acid, a known inducer of tannase, forms insoluble complexes with media proteins, making it difficult to separate pure biomass. Therefore, this study optimizes the production of cell-associated tannase using a "protein-tannin complex" free media. An exploratory study was first conducted in shake-flasks to select the inducer, carbon source, and nitrogen sources. As a result it was found that gallic acid induces tannase synthesis, a tryptose broth gives higher biomass, and lactose supplementation is beneficial. The medium was then optimized using response surface methodology based on the full factorial central composite design in a 3 l bioreactor. A $2^3$ factorial design augmented by 7 axial points (${\alpha}$ = 1.682) and 2 replicates at the center point was implemented in 17 experiments. A mathematical model was also developed to show the effect of each medium component and their interactions on the production of cell-associated tannase. The validity of the proposed model was verified, and the optimized medium was shown to produce maximum cell-associated tannase activity of 9.65 U/l, which is 93.8% higher than the activity in the basal medium, after 12 h at pH 5.0, $30^{\circ}C$. The optimum medium consists of 38 g/l lactose, 50 g/l tryptose, and 2.8 g/l gallic acid.

• Bulletin of the Korean Chemical Society
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• 제33권10호
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• pp.3378-3382
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• 2012

A superhydrophobic surface with a water contact angle > $150^{\circ}$ has attracted great interest from both fundamental and practical aspects. In this study, we demonstrated that hydrophobicity of a silica nanotube (SNT) array can be easily controlled by the SNT aspect ratio. In addition, the adhesive and anti-adhesive properties were controlled without modifying the hydrophobic surface. Various silica structures on a polydimethylsiloxane substrate were prepared using the desired alumina template. Bundle-arrayed and bowl-arrayed silica surfaces exhibited extraordinary superhydrophobicity due to the large frontal surface area and hierarchical micro/nanostructure. As the strategy used in this study is biocompatible and a wide range of hydrophobicities are capable of being controlled by the SNT aspect ratio, a hydrophobic surface composed of an SNT array could be an attractive candidate for bioapplications, such as cell and protein chips.

• 한국식품영양학회지
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• 제6권3호
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• pp.178-188
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• 1993

This research was attempted to investigate changes in physicochemical properties of rice extrudate with added ginger powder extruded by single screw extruder. Graphic three dimensional analysis on response surface regression was used to evaluate effects of extrusion variables on quality factors of the extrudate according to two independent variables, ginger consent 0∼12%, moisture content 14∼26%. The summarized results are as follows : 1) Regarding proximate composition of rice extrudate with added ginger powder, as ginger powder content of raw material Increased, crude tat, crude protein, crude ash and crude fiber increased, while soluble nitrogen free extract decreased. 2) Graphic three dimensional analysis on response surface regression was conducted for each dependent variable which revealed statistically significant relationship with independent variables, 0∼120A ginger and 14∼26% moisture content. Expansion ratio had a critical point as moisture content changed. As ginger and moisture content Increased, bulk density, break strength and water absorption Index Increased, while water solubility Index decreased. The predicted maximum degree of gelatinization in 6.15% ginger and 15.56% moisture content is 88.27%, and lightness decreased as ginger content Increased. According to the microstructure for the cross section of extrudate obsorbed with image analyzer, air cell number and perimeter revealed saddle point, meanwhile total area and fractarea of air cell had critical points as moisture content changed. In view of the results, quality of rice extrudate with added ginger powder was optimum when rice flour was fed to the extruder with 2∼7% singer powder and 15∼20% moisture content.

• Molecular & Cellular Toxicology
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• 제1권2호
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• pp.116-123
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• 2005

The application of high pressure on cellular morphology, proliferation and protein expression of Jurkat cells (human T lymphocyte cell line) has been extensively investigated. In the present study, we manufactured a novel pressure chamber that modulates 5% $CO_{2}$, temperature and pressure (up to 3 ATA). Jurkat cells was incubated 2 ATA pressure and analyzed cellular morphology and growth using an electron microscopy and MTT assay. The cells showed the morphological changes in the cell surface, which appeared to cause a severe damage in cell membrane. The growth rate of the cells under 2 ATA pressure decreased as cultured time got increased. Furthermore, a long term exposure of high pressure on Jurkat cells may act as one of the important cellular stresses that leads to inducing cell death. Cellular proteomes were separated by 2-dimensional electrophoresis with pH 3-10 ranges of IPG Dry strips. And many proteins showed significant up-and-down expressions with hyperbaric pressure. Out of all, 10 spots were identified significantly using matrix-assisted laser desorption/ionization-time of fight (MALDI-TOF) mass spectrometry. We and found that 9 protein expressions were decreased and one protein, heat shock protein HSP 60, was increased in Jurkat cells under 2 ATA. Identified proteins were related to lipid metabolism and signal transduction.

• 미생물학회지
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• 제42권3호
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• pp.195-198
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• 2006

Anthrax lethal toxin은 탄저병의 치사원인이 되는 독소이며, Lethal toxin은 두 종류의 단백질 PA (Protective antigen)과 LF (lethal factor)로 구성되어 진다. PA는 세포표면의 수용체와 결합하여 LF를 세포질 안으로 이동시켜 주는 역할을 한다. LF는 금속 이온$(Zn^{2+})$ 의존적 단백질 가수분해 효소로써 MKKs[MAPK (mitogen-activated protein kinase) kinases] 집단 단백질의 아미노 말단 부분을 절단하여 대상 세포를 죽음으로 유도하는 것으로 알려져 있다. 본 연구에서는 LF에 대한 특성 분석 및 억제제 개발에 과한 연구를 위해 cell-based high-throughtput screens 개발에 선행되어야 하는 기초 자료를 마련하는데 그 목적이 있다. 이를 위하여 LF의 절단 대상이 되는 기질이 MEK1을 yeast내에서 동시 발현시켜 LF의 활성을 검증하였다. 먼저 효모(Saccharomyces cerevisiae)를 숙주로 하여 LF의 기질인 MEK1 발현 vector를 구축하였고, 구축된 발현 system을 기본을 LF 활성을 검증하고자 yeast에 형질전환하여 plasmid의 안전성 및 MEK1 유전자의 발현 및 LF에 의한 MEK1 아미노말단의 절단 부위를 확인하였다. 본 연구는 세포내 검증 system 도입의 기초적 자료를 제공하였으며, yeast내의 MEK1 발현은 탄저병의 저해제 선별 및 활성 측정 검증을 생체에서 고효율적이며, 안정적으로 할 수 있다는 가능성을 나타냈다.

• BMB Reports
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• 제45권8호
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• pp.427-432
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• 2012

Primary cilia, single hair-like appendage on the surface of the most mammalian cells, were once considered to be vestigial cellular organelles for a past century because of their tiny structure and unknown function. Although they lack ancestral motility function of cilia or flagella, they share common ground with multiciliated motile cilia and flagella on internal structure such as microtubule based nine outer doublets nucleated from the base of mother centrioles called basal body. Making cilia, ciliogenesis, in cells depends on the cell cycle stage due to reuse of centrioles for cell division forming mitotic spindle pole (M phase) and assembling cilia from basal body (starting G1 phase and maintaining most of interphase). Ciliary assembly required two conflicting processes such as assembly and disassembly and balance between these two processes determines the length of cilia. Both process required highly conserved transport system to supply needed substance to grow tip of cilia and bring ciliary turnover product back to the base of cilia using motor protein, kinesin and dynein, and transport protein complex, IFT particles. Disruption of ciliary structure or function causes multiple human disorder called ciliopathies affecting disease of diverse ciliated tissues ranging from eye, kidney, respiratory tract and brain. Recent explosion of research on the primary cilia and their involvement on animal development and disease attracts scientific interest on how extensively the function of cilia related to specific cell physiology and signaling pathway. In this review, I introduce general features of primary cilia and recent progress in understanding of the ciliary length control and signaling pathways transduced through primary cilia in vertebrates.

• 한국식품위생안전성학회지
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• 제13권3호
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• pp.206-212
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• 1998

Salmonella spp.의 신속한 검출을 위하여 엷은 박막형태의 수정결정을 사용하는 압전류적(piesoelectric) 항체센서 시스템을 개발하고 증류수, 완충용액, 식염용액 등의 여러 매질 중에서 보여주는 진동 특성을 검토하였다. Salmonella spp. 균 구조항원(Common structural antigen)에 대한 항체를 수정결정에 PEI pre-coating, BSA 가교화, 3-APTES silanizaition, protein A와 DTBP thiolation의 5가지 방법에 의해 고정화한 후 항체 센서의 안정성을 살펴보았다. Salmonella 균을 주입하였을 때 Salmonella 균과 수정 결정에 고정화한 항체와의 결함반응에 의해 수정결정의 질량증가와 이에 따른 진동수 감소가 나타났다. 고정화방법 중 protein A와 DTBP를 이용하여 고정화하는 방법이 센서반응을 가장 안정적이고 재현성 있게 나타내줌을 알 수 있었다. $7.45{\times}10^{7}\;CFU/ml$의 Salmonella 균을 반응 cell 내에 주입하였을 때 protein A를 이용한 고정화의 경우 80Hz, DTBP를 이용한 고정화의 경우 283 Hz의 진동수 감소가 나타났으며, 압전류적 항체센서를 이용할 경우 40분 이내에 Salmonella spp.의 검출이 가능하였다.