• 미생물학회지
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• 제36권3호
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• pp.181-186
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• 2000

Clostridium thermocellum이 생성하는 섬유소분해 효소복합체인 cellulosome은 26개 의 서로 다른 단백질로 구성되어 있으며 그 구성물질로서 calcium을 포함하고 있다. 견고한 구조의 이 복합체에서 구성단백질을 분리하여 그 기능을 연구할 목적으로 이 복합체를 해체 (dissociation)하려 시도하였다. 이 복합체는 calcium을 제거하였을 대 해체되었다. 해체된 구성 단백질들은 MonoQ column chromatogrphy에 의해 구조단백질인 CipA를 포함한 분획, 91 kDa(CelK-tr), 60 kDa 과 57 kDa 단백질로 구성된 분획과 주로 46 kDa(CelA-tr), 또는 71 kDa(CelS-tr) 단백질을 포함하는 분획들로 크게 분리되었다. 대부분의 분획들은 crystalline cellulose 분해 활성을 보였다. 순수 분리된 71 kDa 단백질은 $60^{\circ}C$~$70^{\circ}C$에서 섬 유소 분해시 calcium에 의존적이었으나 46 kDa 단백질은 그렇지 않았다. 46 kDa 단백질은 cellodextrin을 celloviose 및 cellotriose 단위로 절단하며 cellotetraose 로부터 glucose가 생 성되는 것이 관찰되었다. Cellulosome의 섬유소분해 최성 산물이 cellobiose인 것을 고려할 때 개개 구성단백질의 활성이 이 효소 복합체내에서 조절되어 있음을 알 수 있다.

• 대한수의학회지
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• 제35권3호
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• pp.447-458
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• 1995

This study was carried out to characterize nonadrenergic, noncholinergic(NANC) relaxation of porcine retractor penis(PRP) muscle induced by electrical field stimulation(EFS) and to investigate the actions of niric oxide(NO) and vasoactive intestinal polypeptide(VIP) as candidates for NANC neurotransmitters. Biphasic relaxations of PRP muscle were induced by EFS to NANC nerve. Rapid-phase relaxation was observed at low frequency(0.5-16Hz) and slow-phase relaxation followed during high frequency(8-60Hz). Both relaxations were frequency-dependent and TTX($1{\times}10^{-6}M$)-sensitive. L-NAME($2{\times}10^{-5}M$) inhibited the rapid-phase relaxation, but not the slow-phase relaxation. The inhibition of the rapid-phase relaxation with L-NAME was reversed by L-arginine ($1{\times}10^{-3}M$) but not by D-arginine($1{\times}10^{-3}M$). Methylene blue($4{\times}10^{-5}M$) reduced the rapid-phase relaxation. Exogenous No(ExoNO, $1{\times}10^{-5}-1{\times}10^{-4}M$) induced dose-dependent relaxations of PRP muscle. Oxyhemoglobin($5{\times}1^{-5}M$) blocked the relaxation induced by ExoNO and inhibited EFS-induced relaxation. Hydroquinone($1{\times}10^{-4}M$) also abolished the relaxation induced by ExoNO, but did not affect EFS-induced relaxation. L-NAME resistant slow-phase relaxation to EFS was inhibited by ${\alpha}$-chymotrypsin(2.5 U/ml). Both methylene blue($4{\times}10^{-5}M$) and Nethylmaleimide($1{\times}10^{-4}M$) reduced the slow-phase relaxation by EFS. [4-Cl-D-$Phe^6$, $Leu^{17}$]-VIP($3{\times}10^{-6}M$) inhibited the slow-phase relaxation by EFS. External applications of VIP ($1{\times}10^{-7}M$) caused relaxations that were simillar to the L-NAME resistant slow-phase relaxations induced by EFS, and relaxant effects of exogenous VIP were blocked by ${\alpha}$-chymotrypsin(2.5 U/ml).

• BMB Reports
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• 제34권1호
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• pp.33-38
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• 2001

The human PTK6 (also known as Brk) polypeptide, which is deduced from its full-length cDNA, represents a non-receptor protein tyrosine kinase (PTK). It contains SH3, SH2, and tyrosine kinase catalytic domains that are closely related to Src family members. We generated an antihuman PTK6 antibody by immunizing rabbits with a PTK6-specific oligopeptide conjugated to BSA, which corresponds to 11 amino acid residues near the C-terminus. An immunoblot analysis with the antibody detected an expected 52-kDa band in various mammalian transformed cell lines. Immunoprecipitation and immunoblot analyses demonstrated that PTK6 is phosphorylated on the tyrosine residues) and interacts with approximately a 23-kDa tyrosine-phosphorylated polypeptide (most likely a substrate of PTK6) in breast carcinoma T-47D cells. An immunofluorescence analysis demonstrated that PTK6 is localized throughout the cytoplasm of T-47D cells. These results support a possible role for PTK6 in the intracellular signal transduction through tyrosine phosphorylation.

• 대한의생명과학회지
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• 제19권1호
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• pp.41-47
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• 2013

Pluripotent stem cells (PSCs) have lots of potential in biomedical sciences owing to its potential to differentiate into any kind of cells in the body. However, it is still a challenge to culture PSCs on a large scale for application to regenerative medicine. Herein, we introduce a synthetic polymer that enables large-scale suspension culture of human PSCs. By employing suspension culture, it became unnecessary to use conventional substrata such as mouse embryonic fibroblast (MEF) or Matrigel$^{TM}$, which are believed to be main causative sources of xenogeneic contamination in cultured human PSCs in vitro. Human PSCs were cultured in the medium in which elastin-like polypeptide (ELP) dissolved. The ELP in the medium became harden as temperature increases by transforming the medium into a semi-solid gel that supported growth of human PSCs in suspension. Gel-sol transition temperature of ELP can be adjusted by modifying the peptide sequence in which 5 amino acids, Val-Pro-Gly-Xaa-Gly, repeated sequentially. We constructed 3D suspension media having transition temperature around $33{\sim}35^{\circ}C$ using an ELP consisted of 40, 60, or 80 repeats of a monomer, which was Val-Pro-Gly-Val-Gly. Among the ELPs, ELP80 was chosen as the best ELP to support growth of human PSCs in suspension culture. This result suggests that the ELP80 can be a medium component for culturing human PSCs in large-scale.

• Archives of Pharmacal Research
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• 제23권2호
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• pp.187-195
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• 2000

Nucleotide sequence extending 2,3-dihydroxybiphenyl 1,2-dioxygenase gene (pcbC) and 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate hydrolase gene (pcbD) of Pseudomonas sp. DJ-12 was previously analyzed and the two genes were present in the order of pcbD-pcbC preceded by a promoter from Pseudomonas sp. DJ-12. In this study, a 3.8-kb nucleotide sequence located downstream of the pcbC gene was analyzed to have three open reading frames (ORFs) that are designated as orf1, pcbE and orf2 genes. All of the ORFs were preceded by each ribosome-binding sequence of 5-GGAXA-3 (X=G or A). However, no promoter-like sequence and transcription terminator sequence were found in the analyzed region, downstream of pcbC gene. Therefore, the gene cluster appeared to be present in the order of pcbD-pcbC-orf1-pcbE-orf2 as an operon, which is unique organization characterized so far in biphenyl- and PCB-degrading bacteria. The orf1 gene was composed of 1,224 base pairs which can encode a polypeptide of molecular weight 44,950 containing 405 amino acid residues. A deduced amino acid sequence of the orf1 gene product exhibited 21-33％ identity with those of indole dioxygenase and phenol hydroxylase components. The pcbE gene was composed of 783 base pairs encoding 2-hydroxypenta-2,4-dienoate hydratase involved in the 4-chlorobiphenyl catabolism. The orf2 gene was composed of 1,017 base pairs encoding a polypeptide of molecular weight 37,378 containing 338 amino acid residues. A deduced amino acid sequence of the orf2 gene product exhibited 31％ identity with that of a nitrilotriacetate monooxygenase component.

• BMB Reports
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• 제30권5호
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• pp.356-361
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• 1997

We cloned a cDNA (pPRC-1) which was comprised of 841 nucleotides from the cDNA library of a male ICR mouse submandibular gland ($SMG^+$). The nucleotide sequences of pPRC-1 were identical to those of exons 2 and 3 of the mGK-21 gene, a potentially functional glandular kallikrein identified in a Balb/c mouse, except for one nucleotide residue. Although this substitution changes Ile (ATT) in pPRC-1 to Val (GTT) in mGK-21, this difference has been explained by strain polymorphism. From the amino acid sequences predicted from its cDNA, we speculated that mGK-21 gene products/pGK21 consist of 261 amino acids including the $NH_2$-terminal signal peptide (residues 1~17), the short propeptide (residues 17~24), and the active peptide (residues 25~261). Although we did not demonstrate the enzyme activity of pGK21, it was assumed that pGK 21 was involved in the maturation of certain bioactive polypeptide(s) in mouse SMG for the following reasons : (a) mGK-21 gene was apparently expressed in a male ICR mouse SMG: (b) the proposed active site $His^{65}$, $Asp^{120}$, and $Ser^{213}$ residues were completely conserved in pGK21 just like other glandular kallikreins; (c) the cloned cDNA was translated to a predicted 27 kDa polypeptide chain in vitro: (d) the 27 kDa polypeptide chain produced by CHO cells was produced to a putative active form by trypsin.

• 한국잡초학회지
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• 제12권4호
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• pp.368-373
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• 1992

귀리에 alachlor를 처리(處理)할 때 변화(變化)되는 단백질(蛋白質)을 분석(分析)한 결과요약(結果要約)하면 다음과 같다. Alachlor 처리시(處理時) 단백질(蛋白質) 합성(合成)은 $1{\times}10^{-4}$M에서 5.8%, $1{\times}10^{-3}$M에서는 86.5% 억제(抑制)되는 현상(現象)을 보였지만 $1{\times}10^{-5}$M 이하(以下)의 농도(濃度)에서는 증가(增加)되는 현상(現象)을 보였다. 귀리의 근단분열조직(根端分裂組織)에서 추출(抽出)된 단백질(蛋白質)은 100 kd 이하(以下)의 polypeptide로 구성(構成)되어 있으며, alachlor 처리시(處理時) 47kd 이상(以上)의 고분자(高分子) 단백질(蛋白質)들은 억제(抑制)되었지만 23kd 이하(以下)의 저분자(低分子) 단백질(蛋白質)들은 오히려 증가(增加)되는 현상(現象)을 보였다. 이차원적(二次元的) 전기영동(電氣泳動) 결과(結果) alachlor 처리시(處理時) 83kd의 1, 2 spot, 70kd의 3, 4 spot, 47.5kd의 5, 6 spot의 polypeptide가 억제(抑制)된 반면(反面), 20kd의 7 spot, 16kd 의 8, 9, 10 spot는 증가(增加)되는 현상(現象)을 보였다. 또한 귀리 근단(根端) 분열조직(分裂組織)은 산성(酸性) 단백질(蛋白質)로서, 주(主)로 중성(中性) 부위(部位)에 큰 polypeptide spots들이 존재(存在)하고 있으며, 작은 spots을은 산성(酸性)쪽에 분리(分離)되어 나타났다.

• BMB Reports
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• 제33권3호
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• pp.256-262
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• 2000

In this study the disassembly process of chlorophyII (ChI)protein complexes of a stay-green mutant (ore10 of Arabidopsis thaliana) was investigated during the dark incubation of detached leaves. During this dark-induced senescence (DIS), the Chi loss was delayed in the mutant, while the photochemical efficiency of photosystem II (PSII) or Fv/Fm was accelerated when compared with the wild type (WT) leaves. This indicates that the decrease in Fv/Fm is a separate process and not causally-linked to the degradation of Chi during DIS of Arabidopsis leaves. In the native green gel electrophoresis of the Chi-protein complexes, which was combined with an additional twodimensional SDS-PAGE analysis, the delayed senescence of this mutant was characterized by the appearance of an aggregate at 1 d or 2 d, as well as very stable light harvesting complex II (LHCII) trimers until 5 d after the start of DIS. The polypeptide composition of the aggregates varied during the whole DIS at 5 d. Dl protein appeared to be missing in the aggregates. This result supports the idea of a faster depletion of functional PSH in the mutants compared with WT, as suggested by the earlier reduction of Fv/Fm and the stable Chl a/b ratio in the mutants. At 5 d, the WT leaves also often showed aggregates, but the polypeptide composition was different from those of ore10. The results presented suggest that the formation of aggregates, or stable LHCII trimers in the stay-green mutants, is a way to structurally protect Chi-protein complexes from serious proteolytic degradation. Detailed disassembly processes of Chi-protein complexes in WT and ore10 mutants are discussed.

• BMB Reports
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• 제40권5호
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• pp.644-648
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• 2007

Exposure of algae or plants to irradiance from above the light saturation point of photosynthesis is known as high light stress. This high light stress induces various responses including photoinhibition of the photosynthetic apparatus. The degree of photoinhibition could be clearly determined by measuring the parameters such as absorption and fluorescence of chromoproteins. In cyanobacteria and red algae, most of the photosystem (PS) II associated light harvesting is performed by a membrane attached complex called the phycobilisome (PBS). The effects of high intensity light (1000-4000 ${\mu}mol$ photons $m^{-2}s^{-1}$) on excitation energy transfer from PBSs to PS II in a cyanobacterium Spirulina platensis were studied by measuring room temperature PC fluorescence emission spectra. High light (3000 ${\mu}mol$ photons $m^{-2}s^{-1}$) stress had a significant effect on PC fluorescence emission spectra. On the other hand, light stress induced an increase in the ratio of PC fluorescence intensity of PBS indicating that light stress inhibits excitation energy transfer from PBS to PS II. The high light treatment to 3000 ${\mu}mol$ photons $m^{-2}s^{-1}$ caused disappearance of 31.5 kDa linker polypeptide which is known to link PC discs together. In addition we observed the similar decrease in the other polypeptide contents. Our data concludes that the Spirulina cells upon light treatment causes alterations in the phycobiliproteins (PBPs) and affects the energy transfer process within the PBSs.

• 한국잡초학회지
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• 제9권3호
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• pp.245-249
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• 1989

식물생장(植物生長) 억제제(抑制劑)인 butachlor의 작용기구(作用機構)를 구명(究明)하기 위하여 본(本) 제초제(除草劑)가 생장기본요소(生長基本要素)인 세포분열(細胞分裂) 및 단백질(蛋白質) 합성(合成) 억제(抑制)에 미치는 영향(影響)은 다음과 같다. 1. Butachlor의 세포분열(細胞分裂) 억제(抑制) 효과(效果)는 18시간(時間) 처리후(處理後) $10^{-6}M$은 9.3%, $10^{-5}M$은 28.2%의 억제(抑制)를 보였으며, 특히 $10^{-3}M$ 농도(濃度)에서는 81.6%의 현저한 억제(抑制)를 보임으로서 본(本) 제초제(除草劑)는 농도(濃度)가 높아질수록 세포분열(細胞分裂) 억제효과(抑制效果)도 증가(增加)하였다. 2. 단백질(蛋白質) 억제(抑制) 효과(效果)는 18시간(時間) 처리후(處理後) $10^{-4}M$ 농도(濃度)에서 22.9%, $10^{-3}M$은 34.1%의 억제효과(抑制效果)를 보였다. 또한 24시간(時間) 처리후(處理時) $10^{-4}M$은 34.3%로 로서 18시간(時間) 처리후(處理後)의 $10^{-3}M$과 거의 같은 억제효과(抑制效果)를 보이고 있다. 이와같이 butachlor 는 농도(濃度)와 처리시간(處理時間)이 길어짐에 따라서 단백질(蛋白質) 농도(濃度)의 증가(增加)를 보였다. 3. Butachlor 처리(處理)는 전체적인 단백질(蛋白質) 합성(合成)을 억제(抑制)하였고 특히 분자량 16, 18, 30, 43 그리고 43.5KD의 단백질(蛋白質)을 억제(抑制)하는 것으로 나타났으며 귀리(Avena sativa L.)의 root tip은 100,000 이하의 polypeptide subunit의 단백질로 구성(構成)되었다.