• 대한약리학회지
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• 제32권2호
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• pp.243-257
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• 1996

CCK and cholinergic agonist stimulate enzyme release from the pancreatic acini via G-protein-mediated activation of phospholipase C, In contrast secretin and related peptides increase the level of cAMP and activate cAMP-dependent protein kinase. Camostat, a synthetic protease inhibitor, causes pancreatic hypertrophy and hyperplasia by increasing the CCK release. In this study, the secretagogue-induced changes of intracellular proteins were examined in the dispersed pancreatic acini of rats with or without camostat treatment. Camostat(FOY-305, 200 mg/kg, p.o.) was given for 4 days twice daily and the dispersed acini were prepared at 12 bouts after last treatment. The profiles of Intracellular phosphoproteins were analyzed by two-dimensional gel electrophoresis after incubating the acini with $^{32}P$. The amylase release from the dispersed acini was measured. The pancreatic weight was increased to 126% of control, while amylase activity per mg acinar protein decreased to 41% of control, The maximum response of amylase release from dispersed acini to CCK-8 or carbachol was markedly decreased(65% or 46% of control, respectively). The group of intracellular proteins(24 kD, pI $4.5{\sim}8.5$) was increased in quantity by camostat. CCK-8 or secretin increased phosphorylation of a protein(34 kD, pI 4.7) in camostat-treated as well as control rats. CCK-8 increased tyrosine phosphoryiation in the acini of control rats. However, in camostat-treated rats, the basal level of tyrosine phosphorylation was increased and it was rather decreased by CCK-8. Secretin had no effect on the level of tyrosine phosphorylation in acini. These results indicate that both phospholipase C and adenylate cyclase induce phosphorylation of an intracellular acinar protein(34 kD, pI 4.7) and camostat treatment increases the basal level of tyrosine phosphorylation in acinar cells. And these results suggest that not only serine/threonine protein kinase but also protein tyrosine kinase/phosphatase are involved in the process of CCK receptor mediated stimulation-secrelion coupling.

• Parasites, Hosts and Diseases
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• 제59권6호
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• pp.615-623
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• 2021

Human sparganosis is a food-borne parasitic disease caused by the plerocercoids of Spirometra species. Clinical diagnosis of sparganosis is crucial for effective treatment, thus it is important to identify sensitive and specific antigens of plerocercoids. The aim of the current study was to identify and characterize the immunogenic proteins of Spirometra erinaceieuropaei plerocercoids that were recognized by patient sera. Crude soluble extract of the plerocercoids were separated using 2-dimensional gel electrophoresis coupled with immunoblot and mass spectrometry analysis. Based on immunoblotting patterns and mass spectrometry results, 8 antigenic proteins were identified from the plerocercoid. Among the proteins, cysteine protease protein might be developed as an antigen for diagnosis of sparganosis.

• 한국작물학회지
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• 제67권4호
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• pp.211-221
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• 2022

논 토양 조건에서 팥의 생육기 중 초기 생육 단계에 해당되는 유묘기에 과습 스트레스 처리 후 생육특성 변화 및 팥잎의 단백질 발현 양상을 조사한 결과는 다음과 같다. 1. 아라리(밀양 8호)를 공시 품종으로 과습 스트레스 실험을 진행하였다. 3일간 과습 처리하는 동안 팥의 초장과 생체중(잎, 줄기)은 통계적으로 유의하지는 않았으나 생체중(뿌리)와 SPAD value 값은 통계적으로 유의한 결과가 나왔다. 2. 과습 처리 3일 후 잎의 단백질 발현 양상을 확인한 결과 21개의 차별 발현된 단백질 중 과습 스트레스 처리에 의해 9개의 단백질들이 up-regulated 되었고, 12개의 단백질들이 down-regulated 되었다. 단백질을 기능별로 분류한 결과 Biological Process에서 carbohydrate metabolic process와 관련된 단백질들이 가장 많이 나왔고 Cellular Component에서는 Chloroplast에서 가장 많이 존재하였다. 3. Regulatory protein과 관련된 Maturase K 단백질은 처리구에서 대조구보다 발현양이 증가한 것으로 나타났다. 4. Carbohydrate metabolic process와 관련된 Malate dehydrogenase 단백질과 Glyceraldehyde-3-phosphate dehydrogenase 단백질은 과습 스트레스를 받았을 경우 단백질 발현양이 증가하였다. 5. Photosynthesis와 관련된 Carbonic anhydrase (CA)는 처리구보다 대조구에서 단백질 발현량이 더 증가한 것을 확인할 수 있었다. 스트레스와 관련된 단백질 Superoxide dismutase는 처리구에서 대조구보다 단백질 발현양이 증가한 것을 확인할 수 있었다.

• 생명과학회지
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• 제25권7호
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• pp.826-832
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• 2015

본 연구는 인삼의 잎과 뿌리 단백질 추출물에서 활성탄을 이용하여 염, 계면활성제, 색소를 제거하여 단백질체 분석 연구에 미치는 잠재적인 효과에 대한 평가를 기술하고 있다. 5%(w/v) 활성탄(100-400 mesh)과 함께 단백질 추출물을 30분간 4℃에서 반응시켜 염과 계면활성제를 제거한 후 SDS-PAGE를 분석하여 단백질의 양상을 관찰하였다. 엽록소 함량의 분석은 활성탄 처리 후 엽록소의 상당한 양(~33%)이 제거되는 것을 보여주었고, 이 분석은염, 계면활성제 제거만이 아닌 색소의 제거에서도 활성탄의 잠재적 효과가 있음을 보여 주고 있다. 활성탄을 처리한 단백질 시료를 이용하여 이차원 전기영동과 PCA 통계분석을 시행한 결과 단백질은 gel에서 더 나은 해상도를 보여주었으며 단백질 시료의 정제에서도 활성탄의 효과가 있음을 확인하였다. 종합적으로, 이 결과들은 활성탄을 이용한 간단한 방법으로 다양한 식물 조직의 단백질 추출물에서 염, 계면활성제, 색소를 제거함으로써 고해상도의 단백질체 분석에 적용될 수 있을 것으로 기대된다.

• Toxicological Research
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• 제8권2호
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• pp.343-357
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• 1992

Tumor necrosis factor-${\alpha}$(TNF), a polypeptide hormone secreted primarily by activated macrophages, was originally identified on the basis of its ability to cause hemorrhagic necrosis and tumor regression in vivo. Subsequently, TNF has been shown to be an important component of the host responses to infection and cancer and may mediate the wasting syndrome known as cachexia. These systemic actions of TNF are reflected in its diverse effects on target cells in vitro. TNF initiates its diverse cellular actions by binding to specific cell surface receptors. Although TNF receptors have been identified on most of animal cells, regulation of these receptors and the mechanisms which transduce TNF receptor binding into cellular responses are not well understood. Therefore, in the present study, the mechanisms how TNF receptors are being regulated and how TNF receptor binding is being transduced into cellular responses were investigated in rat liver plasma membranes (PM) and ME-180 human cervical carcinoma cell lines. $^{125}I$-TNF bound to high ($K_d=1.51{\pm}0.35nM$)affinity receptors in rat liver PM. Solubilization of PM with 1% Triton X-100 increased both high affinity (from $0.33{\pm}0.04\;to\;1.67{\pm}0.05$ pmoles/mg protein) and low affinity (from $1.92{\pm}0.16\;to\;7.57{\pm}0.50$ pmoles/mg protein) TNF binding without affecting the affinities for TNF, suggesting the presence of a large latent pool of TNF receptors. Affinity labeling of receptors whether from PM or solubilized PM resulted in cross-linking of $^{125}I$-TNF into $M_r$ 130 kDa, 90 kDa and 66kDa complexes. Thus, the properties of the latent TNF receptors were similar to those initially accessible to TNF. To determine if exposure of latent receptors is regulated by TNF, $^{125}I$-TNF binding to control and TNF-pretreated membranes were assayed. Specific binding was increased by pretreatment with TNF (P<0.05), demonstrating that hepatic PM contains latent TNF receptors whose exposure is promoted by TNF. Homologous up-regulation of TNF receptors may, in part, be responsible for sustained hepatic responsiveness during chronic exposure to TNF. As a next step, the post-receptor events induced by TNF were examined. Although the signal transduction pathways for TNF have not been delineated clearly, the actions of many other hormones are mediated by the reversible phosphorylation of specific enzymes or target proteins. The present study demonstrated that TNF induces phosphorylation of 28 kDa protein (p28). Two dimensional soidum dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) resolved the 28kDa phosphoprotein into two isoforms having pIs of 6.2 and 6.1. The pIs and relative molecular weight of p28 were consistent with those of a previously characterized mRNA cap binding protein. mRNA cap binding proteins are a class of translation initiation factors that recognize the 7-methylguanosine cap structure found on the 5' end of eukaryotic mRNAs. In vitro, these proteins are defined by their specific elution from affinity columns composed of 7-methylguanosine 5'-triphosphate($m^7$GTP)-Sepharose. Affinity purification of mRNA cap binding proteins from control and TNF treated ME-180 cells proved that TNF rapidly stimulates phosphorylation of an mRNA cap binding protein. Phosphorylation occurred in several cell types that are important in vitro models of TNF action. The mRNA cap binding protein phosphorylated in response to TNF treatment was purifice, sequenced, and identified as the proto-oncogene product eukaryotic initiation factor-4E(eIF-4E). These data show that phosphorylation of a key component of the cellular translational machinery is a common early event in the diverse cellular actions of TNF.

• 한국작물학회지
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• 제63권1호
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• pp.25-34
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• 2018

휴면성이 다른 백중밀, 금강밀, 우리밀 후숙종자의 ABA 농도에 따른 발아 및 배아에서의 단백질체 발현 특성을 조사한 결과는 다음과 같다. 1. 백중밀, 금강밀, 우리밀 등 3품종의 0, 10, 30 및 $50{\mu}M$ ABA에서의 평균 발아지수와 발아율은 각각 0.95와 98% 이상으로 통계적으로 유의한 차이가 없었다. 유아와 유근의 생장은 $0{\mu}M$ ABA보다 10, 30 및 $50{\mu}M$ ABA에서 생장이 크게 억제되었는데, ABA 농도가 높을수록 생장이 더 억제되었다. 2. 3품종 배아의 평균 ABA 함량은 $0{\mu}M$ ABA와 $50{\mu}M$ ABA에서 각각 0.78 ng/mg과 269.04 ng/mg으로서 농도에 따른 ABA 함량의 차이가 컸다. 3. $0{\mu}M$ ABA 처리구에 비하여 $50{\mu}M$ ABA 처리구에서 발현양이 증가한 단백질 spot (S1, S3, S4, S6, S15, S16, S17)은 7개였으며, 감소한 단백질 spot (S2, S5, S9, S10, S11, S12, S13, S14, S18)은 8개였고, 증가와 감소가 동시에 이루어진 단백질 spot (S2)은 1개였다. $50{\mu}M$ ABA에서만 검출된 단백질 spot (S7, S8)은 2개였다. 4. 각 단백질 spot의 $0{\mu}M$ ABA 처리구 양에 대한 $50{\mu}M$ ABA 처리구 양의 평균 배수 값(fold 값)이 1.5배 이상으로 증가한 단백질 spot은 S1 (globulin-3A), S6 (globulin-1 S allele), S16 (globulin-1 S allele), S17 (globulin-1 S allele) 등으로 모두 globulin류 단백질이었다. 또한 $50{\mu}M$ ABA에서만 확인된 단백질 spot인 S7 (globulin 3)과 S8 (globulin-1 S allele)도 globulin 단백질이었다. 5. 각 단백질 spot의 $0{\mu}M$ ABA 처리구 양에 대한 $50{\mu}M$ ABA 처리구 양의 평균 배수 값(fold 값)이 0.7 이하로 감소한 단백질 spot은 S10 (glutamine sysnthetase cytosolic isozyme), S12 (S-adenosylmethionine synthetase 2), S14 (isocitrate dehydrogenase NADP)이었다. 이상의 결과는 ABA에 의한 밀 유묘의 유아와 유근의 생장억제는 배아에서의 ABA 농도 증가, 그리고 이에 따른 배아의 glutamine 합성에 관여하는 다양한 효소의 발현 감소 및 메틸기공여물질의 감소와 이에 따른 메틸기 전이활성의 감소 등이 관여하고 있음을 의미한다. 한편 배아에서의 ABA에 의한 globulin 단백질의 증가는 배아 특이적 globulin의 일시적 합성 증가와 globulin 분해 효소의 활성 억제 등이 복합적으로 관여한 결과로 생각된다.