• Asian-Australasian Journal of Animal Sciences
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• 제25권11호
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• pp.1540-1545
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• 2012

Steroidogenesis requires coordination of the anabolic and catabolic pathways of lipid metabolism, but the profile of proteins associated with progesterone synthesis in cyclic and pregnant corpus luteum (CL) is not well-known in cattle. In Experiment 1, plasma progesterone level was monitored in cyclic cows (n = 5) and pregnant cows (n = 6; until d-90). A significant decline in the plasma progesterone level occurred at d-19 of cyclic cows. Progesterone level in abbatoir-derived luteal tissues was also determined at d 1 to 5, 6 to 13 and 14 to 20 of cyclic cows, and d-60 and -90 of pregnant cows (n = 5 each). Progesterone level in d-60 CL was not different from those in d 6 to 13 CL and d-90 CL, although the difference between d 6 to 13 and d-90 was significant. In Experiment 2, protein expression pattern in CL at d-90 (n = 4) was compared with that in CL of cyclic cows at d 6 to 13 (n = 5). Significant changes in the level of protein expression were detected in 32 protein spots by two-dimensional polyacrylamide gel electrophoresis (2-DE), and 23 of them were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Six proteins were found only in pregnant CL, while the other 17 proteins were found only in cyclic CL. Among the above 6 proteins, vimentin which is involved in the regulation of post-implantation development was included. Thus, the protein expression pattern in CL was disorientated from cyclic luteal phase to mid pregnancy, and alterations in specific CL protein expression may contribute to the maintenance of pregnancy in Korean native cows.

• 한국작물학회지
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• 제60권1호
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• pp.33-40
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• 2015

재배환경에 따른 단백질 발현을 이차원전기영동을 이용하여 단백질 발현양상을 확인하고 이미지 분석을 통하여 단백질 발현 정도를 수치화하여 비교 분석한 결과, 재배환경에 따른 단백질 발현 양상은 전반적으로 매우 유사하였으며, 주요 단백질의 발현에는 차이가 없었다. 하지만 논과 밭, 즉 재배환경은 단백질의 발현 정도에는 영향을 미치는 것을 확인 할 수 있었다. 공시품종들 중에서 백천콩은 다른 세 품종과는 다르게 논에서 단백질 발현 정도가 밭에서의 단백질 발현 정도보다 높았다. 이 결과로 보아 품종마다 재배환경에 따른 단백질의 발현 정도에 차이가 있는 것으로 사료된다. 향후, 단백질 총 함량을 재배환경에 따라 비교하고 또한 질량분석을 통하여 증가 또는 감소되는 단백질의 기능을 확일 할 필요가 있다고 사료된다.

• 한국응용곤충학회지
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• 제46권2호
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• pp.229-234
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• 2007

호박벌 일벌독의 성분과 생리활성을 규명하기 위하여 단백질 성분분석과 암세포 생육 저해 효능, 항균력을 검토하였다. 이차원단백질 분석을 통해 호박별의 일벌독은 63개의 단백질이 존재하는 것으로 확인되었으며, 가장 많은 함량을 보이는 3개의 단백질을 염기서열 분석하였다. 그러나 이들 성분은 아직 밝혀지지 않은 성분으로 판단되었다. 호박벌 일벌독의 암세포 (간암; Hep3B, 폐암; A549, 유방암; BT-20, 위암; AGS) 에 대한 생육 저해능은 시료 농도가 증가함에 따라 증가하는 경향을 보이며 100ng/ml에서 간암세포 (Hep3B) 에 대한 생육 저해능이 55%로 가장 높았다. 항균활성에 E. faecium 과 S. sonnei에 대하여 최소발육억제농도와 최소살균농도 모두 각각 0.256ug/ml 로 강한 항균활성을 나타내었으며, 그 외의 피검균에 대해서도 비교적 높은 활성을 보였다. 이상의 결과로부터 호박벌 일벌독의 성분은 다른 벌의 독성분과는 차이를 보이며, 그 생리활성에 있어 약리학적 이용이 가능할 것으로 판단되었다.

• Bulletin of the Korean Chemical Society
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• 제34권8호
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• pp.2419-2424
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• 2013

Guanosine-5'-diphosphate 3'-diphosphate (ppGpp) serves as alarmone in bacterial stringent responses. In this study, an affinity column was constructed by immobilizing ppGpp to NHS-Sepharose for isolating ppGpp-binding proteins. A novel ppGpp-binding protein, YjgA, was isolated and characterized by MALDI-TOF MS (matrix-assisted laser desorption ionization-time-of-flight mass spectrometry) coupled with two-dimensional gel electrophoresis. YjgA and truncated forms of YjgA were cloned and over-expressed in BL21 (DE3). The binding affinity of YjgA to ppGpp was determined by equilibrium dialysis. The interaction of YjgA with ppGpp was very specific, considering that the dissociation constant of YjgA with ppGpp was measured as $5.2{\pm}2.0{\mu}M$, while the affinities to GTP and GDP were about 60 and 30 times weaker than ppGpp. Expression of yjgA gene in Escherichia coli K-12 MG1655 was examined by reverse transcription polymerase chain reaction (RT-PCR). RT-PCR results revealed that yjgA was expressed from early to late stationary phase. The yjgA deletion mutant exhibited decreased cell number at stationary phase compared to parent strain and the over-expression of YjgA increased the cell number. These results suggested that YjgA might stimulate cell division under stationary phase. In most prokaryotic genome, about half of the protein candidates are hypothetical, that are expected to be expressed but there is no experimental report on their functions. The approach utilized in this study may serve as an effective mean to probe the functions of hypothetical proteins.

• 한국동물학회지
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• 제33권4호
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• pp.461-467
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• 1990

Acrylamide의 신경독성에 대해 조사하기 위하여 초기 후지마비 증상이 유도된 생쥐의 뇌를 SDS-PAGE와 이차원 전기영동법으로 분석한 결과, SDS-polyacrylamid gel 전기영동에 의해 처리군, 대조군 및 회복군을 비교하여 보았을때 두드러진 양상의 차이를 나타내지 않았으나 이차원 전기영동상에서 실험군간의 다소 상이한 단백질 양상이 관찰되었다. 특히, 처리군에서 spot 20(14,500, 5.64),21(19,900, 6.78)및 수개의 단백질이 소실되었으며 spot 9(31,300, 5.82),11(31,300, 5.36) 및 19(16,400, 5.42)들은 비교적 많이 감소되었다. 처리군에서 양적변화를 보인 이들 spot중에서 spot 20 (14,500, 5.64)는 회복군에서 대조군 보다 현저히 증가하였으나 spot 11(31,300, 5.36),19(16,400, 5.42) 및 다수의 spot들은 회복군에서 대조군과 대등한 양상을 보여주었다. 위와같은 실험결과는 acrylamide가 초기적 말초신경 장애를 나타내는 시기에 이미 뇌의 단백질대사에 관여함으로써 단백질합성이 저해되고, 이로 인하여 뇌의 조절능력이 부진하게 되어 결과적으로 중추신경계의 이상이 일어나게 된다는 것을 시사한다. 또한 손사에 대한 회복은투여된 농도 및 기간에 의존하여 acrylamide에 대한 노출이 중지된 후 점진적인 자구적 복구에 의해 이루어지는 것으로 사료된다.

• Journal of Microbiology and Biotechnology
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• 제25권2호
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• pp.152-161
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• 2015

CodY is a highly conserved protein in low G+C gram-positive bacteria that regulates genes involved in sporulation and stationary-phase adaptation. Bacillus thuringiensis is a grampositive bacterium that forms spores and parasporal crystals during the stationary phase. To our knowledge, the regulatory mechanism of CodY in B. thuringiensis is unknown. To study the function of CodY protein in B. thuringiensis, BMB171codY^- was constructed in a BMB171 strain. A shuttle vector containing the ORF of cry1Ac10 was transformed into BMB171 and BMB171codY^-, named BMB171cry1Ac and BMB171codY^-cry1Ac, respectively. Some morphological and physiological changes of codY mutant BMB171codY^-cry1Ac were observed. A comparative proteomic analysis was conducted for both BMB171codY^-cry1Ac and BMB171cry1Ac through two-dimensional gel electrophoresis and MALDI-TOF-MS/MS analysis. The results showed that the proteins regulated by CodY are involved in microbial metabolism, including branched-chain amino acid metabolism, carbohydrate metabolism, fatty acid metabolism, and energy metabolism. Furthermore, we found CodY to be involved in sporulation, biosynthesis of poly-β-hydroxybutyrate, growth, genetic competence, and translation. According to the analysis of differentially expressed proteins, and physiological characterization of the codY mutant, we performed bacterial one-hybrid and electrophoretic mobility shift assay experiments and confirmed the direct regulation of genes by CodY, specifically those involved in metabolism of branched-chain amino acids, ribosomal recycling factor FRR, and the late competence protein ComER. Our data establish the foundation for in-depth study of the regulation of CodY in B. thuringiensis, and also offer a potential biocatalyst for functions of CodY in other bacteria.

• 한국가금학회:학술대회논문집
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• 한국가금학회 2005년도 제22차 정기총회 및 학술발표회
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• pp.66-67
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• 2005

배자 생식세포 발달에 관련된 메카니즘을 밝혀내기 위해서, 닭 배자 생식기에서 추출한 원시 생식세포의 단백질체 지도를 만들었다. 총 500 배자를 6일간 배양하여 배자 생식기를 획득했고, 7-10일 배양 후, 배양된 원시생식세포는 2차원 젤 전기 영동법에 의해 분할되어 졌다. 유의적 발현 수준을 나타낸 많은 단백질 스팟 들은 MALDI-TOP 와 LC-MS/MS에 의해 확인되었으며, 89개의 단백질 스팟 중에 50개의 mass spectra 들이 데이터베이스에서 조류 단백질과 일치함을 확인하였다. 본 실험에서 행한 단백질체 지도는 형질전환 연구와 생식세포 생물학 분야에 중요한 참고 문헌으로 가치를 가질수 있을 것이다.

• Archives of Pharmacal Research
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• 제29권3호
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• pp.224-234
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• 2006

We employed human SK-MEL-28 cells as a model system to identify cellular proteins that accompany N-(4-methyl)phenyl-O-(4-methoxy)phenyl-thionocarbamate (MMTC)-induced apoptosis based on a proteomic approach. Cell viability tests revealed that SK-MEL-28 skin cancer cells underwent more cell death than normal HaCaT cells in a dose-dependent manner after treatment with MMTC. Two-dimensional electrophoresis in conjunction with matrixassisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry analysis or computer matching with a protein database further revealed that the MMTC-induced apoptosis is accompanied by increased levels of caspase-1, checkpoint suppressor-1, caspase-4, NF-kB inhibitor, AP-2, c-Jun-N-terminal kinase, melanoma inhibitor, granzyme K, G1/S specific cyclin D3, cystein rich protein, Ras-related protein Rab-37 or Ras-related protein Rab-13, and reduced levels of EMS (oncogene), ATP synthase, tyrosine-phosphatase, Cdc25c, 14-3-3 protein or specific structure of nuclear receptor. The migration suppressing effect of MMTC on SK-MEL-28 cell was tested. MMTC suppressed the metastasis of SK-MEL-8 cells. It was also identified that MMTC had little angiogenic effect because it did not suppress the proliferation of HUVEC cell line. These results suggest that MMTC is a novel chemotherapeutic and metastatic agents against the SK-MEL-28 human melanoma cell line.

• 식물조직배양학회지
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• 제21권1호
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• pp.15-22
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• 1994

쎌러리(Apium graveolens L.)의 체세포배 발생 및 인공종자 발아에 있어서 분자수준의 기작을 이해하기 위하여 ABA 및 저온처리에 의한 단백질합성변화에 관한 연구를 수행하였다. 단백질함량과 질산환원효소 활성도는 ABA 및 저온처리한 체세포배 및 유식물은 처리하지 아니한 것에 비하여 특히 유식물에서보다 체세포배 발생시에 더욱 높았다. 2차원 전기영동결과 ABA 및 저온처리에 의하여 심장 형배 시기에서는 30 KD, 32 KD, 171 KD 및 205 KD의 단백질과 자엽시기배에서는 29 KD, 33 KD, 37 KD, 38 KD, 41 KD, 55 KD, 66 KD, 및 110 KD 단백질이 합성되어졌다. 또한 심장형배에서는 42 KD, 44 KD, 59 KD, 64 KD, 101 KD, 104 KD, 및 190 KD의 단백질과 자엽시기에서는 29 KD 및 116 KD의 단백질이 억제되었다. 체세포배 발생 및 인공종자 발아에 있어서 ABA및 저온처리에 의해서 단백질이 합성되어지거나 억제되는 것이 동시에 일어났으며 주로 산성단백질에서 변화가 일어났다. 이와같은 결과는 체세포배 발생과정에서 그리고 체세포배 발아와 유식물의 생장과정에서 환경변화에 적응하기 위한 대사상의 변화가 일어나는 것으로 추정된다.

• Asian Pacific Journal of Cancer Prevention
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• 제15권7호
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• pp.3311-3317
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• 2014

Background: The consequence of Rho GDP dissociation inhibitor alpha (RhoGDI${\alpha}$) activity on migration and invasion of estrogen receptor positive ($ER^+$) and negative ($ER^-$) breast cancer cells has not been studied using the proteomic approach. Changes in expression of RhoGDI${\alpha}$ and other proteins interacting directly or indirectly with RhoGDI${\alpha}$ in MCF7 and MDA-MB-231, with different metastatic potentials is of particular interest. Materials and Methods: $ER^+$ MCF7 and ER- MDA-MB-231 cell lines were subjected to two-dimensional electrophoresis (2-DE) and spots of interest were identified by matrix-assisted laser desorption/ionization time of- flight/time-of-flight (MALDI-TOF/TOF) mass spectrometry (MS) analysis after downregulation of RhoGDI${\alpha}$ using short interfering RNA (siRNA) and upregulated using GFP-tagged ORF clone of RhoGDI${\alpha}$. Results: The results showed a total of 35 proteins that were either up- or down-regulated in these cells. Here we identifed 9 and 15 proteins differentially expressed with silencing of RhoGDI${\alpha}$ in MCF-7 and the MDA-MB-231 cells, respectively. In addition, 10 proteins were differentially expressed in the upregulation of RhoGDI${\alpha}$ in MCF7, while only one protein was identified in the upregulation of RhoGDI${\alpha}$ in MDA-MB-231. Based on the biological functions of these proteins, the results revealed that proteins involved in cell migration are more strongly altered with RhoGDI-${\alpha}$ activity. Although several of these proteins have been previously indicated in tumorigenesis and invasiveness of breast cancer cells, some ohave not been previously reported to be involved in breast cancer migration. Hence, these proteins may serve as useful candidate biomarkers for tumorigenesis and invasiveness of breast cancer cells. Conclusions: Future studies are needed to determine the mechanisms by which these proteins regulate cell migration. The combination of RhoGDI${\alpha}$ with other potential biomarkers may be a more promising approach in the inhibition of breast cancer cell migration.