• BMB Reports
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• 제34권6호
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• pp.517-525
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• 2001

Six renaturable protein kinases that utilize the myelin basic protein (MBP) as a substrate were activated during prolonged exposure of cardiac myocytes to okadaic acid (OA). We characterized the substrate preference and activation of these kinases, with particular emphasis on 3 novel kinases-MBPK-55, MBPK-62 and MBPK-87. The transcription factors c-Jun, Elk, ATF2, and c-Fos that are used to assess mitogen-activated protein kinase activation were all poor substrates for these three kinases. MAPKAPK2 was also not phosphorylated. In contrast, Histone IIIS was phosphorylated by MBPK-55 and MBPK-62. These protein kinases were activated in cultured cardiac fibroblasts, H9c2 cardiac myoblasts, and Cos cells. High concentrations (0.5 to $1\;{\mu}M$) of OA were essential for the activation of the protein kinases in all of the cell types examined, whereas calyculin A [an inhibitor of protein phosphatase 1 (PP1) and PP2A], cyclosporin A (a PP2B inhibitor), and an inactive OA analog all failed to activate these kinases. The high dose of okadaic acid that is required for kinase activation was also required for phosphatase inhibition, as assessed by immunoblotting whole cell lysates with anti-phosphothreonine antibodies. A variety of chemical inhibitors, including PD98059 (MEK-specific), genistein (tyrosine kinase-specific) and Bisindolylmaleimide I (protein kinase C-specific), failed to inhibit the OA activation of these kinases. Thus, MBPK-55 and MBPK-62 are also Histone IIIS kinases that are widely expressed and specifically activated upon exposure to high OA concentrations.

• 한국식물학회:학술대회논문집
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• 한국식물학회 2001년도 춘계학술발표대회:발표눈문요지록
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• pp.25-25
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• 2001

• Natural Product Sciences
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• 제15권1호
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• pp.32-36
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• 2009

Glutamate causes neurotoxicity through formation of reactive oxygen species and activation of mitogen-activated protein kinase (MAPK) pathways. MAPK phosphatase-1 (MKP-1) is one of the phosphatases responsible for dephosphorylation/deactivation of three MAPK families: the extracellular signal-regulated kinase-1/2 (ERK-1/2), the c-Jun N-terminal kinase-1/2 (JNK-1/2), and the p38 MAPK. In this report, the potential involvement of MKP-1 in neuroprotective effects of curcumin, the active ingredient of turmeric (Curcuma longa), was examined using HT22 cells. Glutamate caused cell death and activation of ERK-1/2 but not p38 MAPK or JNK-1/2. Blockage of ERK-1/2 by its inhibitor protected HT22 cells against glutamate-induced toxicity. Curcumin attenuated glutamate-induced cell death and ERK-1/2 activation. Interestingly, curcumin induced MKP-1 activation. In HT22 cells transiently transfected with small interfering RNA against MKP-1, curcumin failed to inhibit glutamate-induced ERK-1/2 activation and to protect HT22 cells from glutamate-induced toxicity. These results suggest that curcumin can attenuate glutamate-induced neurotoxicity by activating MKP-1 which acts as the negative regulator of ERK-1/2. This novel pathway may contribute to and explain at least one of the neuroprotective actions of curcumin.

• 한국동물학회지
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• 제35권3호
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• pp.277-286
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• 1992

개구리의 난자로 부터 maturation promoting factor(MPF)를 추출, 부분 분리하여 이들의 활성을 조사하고 이 물질의 생성과 protein kinase C(반KC)와의 관계를 조사하SB다. 성숙된 난자를 분쇄한 후 초원심분리과정을 거쳐 MPF의 crude extract(CE)를 얻은 다음 ultrafiltration (UF)과 고속액체크로마토그라피를 거쳐서 3종류의 분획 (peak 1, 11, and 111)을 얻었다. 이들 분획을 in nitro assay와 autoradiDgraphy를 사용하여 확인한 결과 분획 11에서 MPF 활성이 있는 것을 알았다. 분리 단계에 따라 MPF의 정제도를 Hl histone kinase assay로 조사한 결깍 UF를 거친 것은 CE보다 약 3배로, 분획 11에서는 약 117배로 증가한 것을 확인하였다. 또한 MPF분획의 인산화를 autoradiography로 조사한 결과 45 KD 단백질을 포함한 수종의 난자 단백질이 강하게 인산화되었음을 알 수 있었다. PKC의 활성화가 난자내 MPF의 생성을 유도하는가를 보기 위하여 PKC의 활성제인 12-0-tetradecanoyl phorbol 13 acetate(TPA)를 처리한 난자의 세포질 추출물을 미세주입 법으로 조사한 결과 TPA 처리 후 6시간부터 난자내 MPF의 활성이 나타나는 것을 알 수 있었다. 이러한 결과들은 PKC의 활성화가 MPF의 생성을 유도하고, MPF의 활성화와 함께 일부 단백질들의 인산화를 통하여 궁극적으로 난자 성숙을 촉진했음을 시사한다.

• Restorative Dentistry and Endodontics
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• 제32권5호
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• pp.459-468
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• 2007

이 연구에서는 법랑모세포 분화과정에서 APin의 기능을 알아보고자 APin-protein microarray를 시행한 후 치아발생과 관련이 있는 MEF2, Aurora kinase A, BMPR-IB와 EF-hand calcium binding protein을 분석하여 다음과 같은 결과를 얻었다. 1 CMV-APin construct를 transfection하여 APin의 과발현을 유도한 경우에는 MEF2와 Aurora kinase A 둘 모두에서 발현이 현저히 감소한 반면에, APin의 발현억제를 유도한 경우에는 둘 모두 변화가 없었다. 2. APin의 과발현을 유도한 경우에는 BMPR-IB와 EF-hand calcium binding protein 모두에서 발현이 크게 증가한 반면, APin을 발현억제 시킨 경우에는 BMPR-IB는 변화가 없었고, EF-hand calcium binding protein은 현저히 감소하였다. 위의 결과들로 보아 APin 단백질은 MEF2, Aurora kinase A, BMPR-IB, EF-hand calcium binding protein과 상호작용하여 법랑모세포의 분화와 석회화 과정 중에 중요한 역할을 하는 것으로 사료된다.

• 한국가축번식학회지
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• 제22권3호
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• pp.265-276
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• 1998

In an attempt to evaluate the function of MAP kinase of porcine oocytes and to develop a method of assessment for kinase activity, we used MBP as a substrate to detect the MAP kinase activity of porcine oocytes matured in in vitro. The MAP kinase which had lower activity during the first 20 hours of culture started to show an increased amount of activity at 25 hours at which a collapse in nuclear membrane was induced. Significant (P<0.05) a, pp.ared at 30 hours of being cultured. The gel phosphorylation method, MBP which has been known to be a substrate for kinase such as cdc2 kinase, was phosphorylated at two positions corresponding to ERK 1 (44kDa) and ERK2 (42 kDa) which are known as mammalian MAP kinase. The existence of MARKK and MAP kinase were identified with western blotting at 0 hour culture of immature GV oocytes. The amount of those proteins did not increase during 40 hours of culture, which suggest that the increase of MAP kinase activity was caused by phosphorylaton rather than due to change in protein amount. MAPKK and MAP kinase were shown to be dephosporylated with deactivated at M 1 stage by inhibition of protein synthesis with cycloheximide added at the strat following the cultrue. We have reulsts that indicate the existedence of MAP kinase cascade which was activated simultaneously with start of porcine oocyte maturation (GVBD).

• Journal of Yeungnam Medical Science
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• 제29권2호
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• pp.77-82
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• 2012

AMP-activated protein kinase (AMPK) is an important cellular fuel sensor. Its activation requires phosphorylation at Thr-172, which resides in the activation loop of the ${\alpha}1$ and ${\alpha}2$ subunits. Several AMPK upstream kinases are capable of phosphorylating AMPK at Thr-172, including LKB1 and CaMKK${\beta}$ ($Ca^{2+}$/calmodulin-dependent protein kinase kinase${\beta}$). AMPK has been implicated in the regulation of physiological signals, such as in the inhibition of cholesterol fatty acid, and protein synthesis, and enhancement of glucose uptake and blood flow. AMPK activation also exhibits several salutary effects on the vascular function and improves vascular abnormalities. AMPK is modulated by numerous hormones and cytokines that regulate the energy balance in the whole body. These hormone and cytokines include leptin, adiponectin, ghrelin, and even thyroid hormones. Moreover, AMPK is activated by several drugs and xenobiotics. Some of these are in being clinically used to treat type 2 diabetes (e.g., metformin and thiazolidinediones), hypertension (e.g., nifedipine and losartan), and impaired blood flow (e.g., aspirin, statins, and cilostazol). I reviewed the precise mechanisms of the AMPK activation pathway and AMPK-modulating drugs.

• 한국미생물·생명공학회지
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• 제21권1호
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• pp.54-58
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• 1993

미생물 대사산물로부터 phorbol ester에 의해 유도되는 K562 세포 표면의 소포형성 및 Protein kinase C(PKC)에 대한 저해제를 탐색하여 방선균 분리주 No.2007-18로 부터 용매추출 및 크로마토그래피의 기법을 이용하여 MT-2007을 분리하였다. MT-2007는 503.9MuM의 농도에서 phorbol 12,13-dibutylate에 의해 유도된 K562 세포표면의 bleb형성을 완전히 저해하였고, PKC 효소의 IC50 값은 31.4 MuM 이었다.

• 대한물리치료과학회지
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• 제11권1호
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• pp.20-27
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• 2004

It is widely accepted that smooth muscle contraction is triggered by intracellular $Ca^{2+}$ ($[Ca^{2+}]_i$) released from intracellular $Ca^{2+}$ stores such as sarcoplasmic reticulum (SR) and from the extracellular space, The increased $[Ca^{2+}]_i$ can phosphorylate the 20-kDa myosin light chain ($MLC_{20}$) by activating MLC kinase (MLCK), and this initiates smooth muscle contraction. In addition to the $[Ca^{2+}]_i$-MLCK-tension pathway, a number of intracellular signal molecules, including mitogen-activated protein kinase (MAPK), protein kinase C (PKC), phosphatidylinositol 3-kinase (PI3K), and Rho-associated coiled coil-forming protein kinase (ROCK), play important roles in the regulation of smooth muscle contraction. However, the mechanisms regulating contraction of caldesmon (CaD), actin-binding protein, are not entirely elucidated in the presence of $Ca^{2+}$. It is known that CaD tightly interacts with actin and inhibits actomyosin ATPase activity. Therefore, the purpose of the present study was to investigate the roles of $Ca^{2+}$-dependent CaD in smooth muscle contraction. Endothelin-1 (ET-1), G-protein coupled receptor agonist and vasoconstrictor, increased both vascular smooth contraction and phosphorylation of CaD in the presence of $Ca^{2+}$. These results suggest that ET-1 induces contraction and phosphorylation of CaD in rat aortic smooth muscle, which may he mediated by the increase of $[Ca^{2+}]_i$.

• Journal of Plant Biotechnology
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• 제43권1호
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• pp.30-36
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• 2016

Brassinosteroids (BRs) are essential plant steroid hormones required for cell elongation, plant growth, development and abiotic and biotic stress tolerance. BRs are recognized by BRI1 receptor kinase that is localized in the plasma membrane, and the BRI1 protein will eventually autophosphorylate in the intracellular domain and transphosphorylate BAK1, which is a co-receptor in Arabidopsis thaliana. However, little is known of the role OsBRI1 receptor kinase plays in Oryza sativa, monocotyledonous plants, compared to that in Arabidopsis thaliana, dicotyledonous plants. As such, we have studied OsBRI1 receptor kinase in vitro and in vivo with recombinant protein and transgenic plants, whose phenotypes were also investigated. A OsBRI1 cytoplasmic domain (CD) recombinant protein was induced in BL21 (DE3) E.coli cells with IPTG, and purified to obtain OsBRI1 recombinant protein. Based on Western blot analysis with phospho-specific pTyr and pThr antibodies, OsBRI1 recombinant protein and OsBRI1-Flag protein were phosphorylated on Threonine residue(s), however, not on Tyrosine residue(s), both in vitro and in vivo. This is particularly intriguing as AtBRI1 protein was phosphorylated on both Ser/Thr and Tyr residues. Also, the OsBRI1 full-length gene was expressed in, and rescued, bri1-5 mutants, such as is seen in normal wild-type plants where AtBRI1-Flag rescues bri1-5 mutant plants. Root growth in seedlings decreased in Ws2, AtBRI1, and 3 independent OsBRI1 transgenic seedlings and had an almost complete lack of response to brassinolide in the bri1-5 mutant. In conclusion, OsBRI1, an orthologous gene of AtBRI1, can mediate normal BR signaling for plant growth and development in Arabidopsis thaliana.