• Title/Summary/Keyword: calmodulin 2

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Inhibition of Calmodulin-Dependent Protein Kinase II by Cyclic and Linear Peptide Alkaloids from Zizyphus Species

  • Han Yong Nam;Hwang Keum Hee;Han Byung Hoon
    • Archives of Pharmacal Research
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    • v.28 no.2
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    • pp.159-163
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    • 2005
  • The effects of sedative peptide alkaloids from Zizyphus species on calmodulin- dependent protein kinase II were investigated. Protein kinase II activity was assayed on the basis of its ability to activate tryptophan 5-monooxygenase as its substrate in the presence of calmodulin. All thirteen alkaloids tested were stronger inhibitors than chlorpromazine ($IC_50,\;98{\mu}M$) on calmodulin-dependent protein kinase II. Among them, the most potent inhibitor was daechuine S27 ($IC_{50},\;2.95{\mu}M$), which was stronger than pimozide ($IC_{50},\;15.0{\mu}M$).

Calmodulin of Olive Flounder Paralichthys olivaceus : Cloning and Expression Analysis

  • Hong, Gyeong-Eun;Kong, Hee Jeong;Nam, Bo-Hye;Kim, Young-Ok;Kim, Woo-Jin;Lee, Sang-Jun;Choi, Tae-Jin
    • Journal of Marine Bioscience and Biotechnology
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    • v.2 no.4
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    • pp.234-237
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    • 2007
  • Calmodulin (CaM) is a $Ca^{2+}$-binding protein essential for biological functions mediated through $Ca^{2+}$-dependent mechanism. A cDNA clone for CaM was isolated from a cDNA library of olive flounder Paralichthys olivaceus. The CaM cDNA concists of 782 bp and encodes a polypeptide of 149 amino acids with four $Ca^{2+}$-binding motifs EF-hands (EF-I, EF-II, EF-III, and EF-IV). The deduced amino acid sequence of CaM shows 97-100% amino acid sequence identity to other CaM sequences. Semi-quantitative PCR analysis revealed that the CaM transcription was began during early development and the CaM mRNA is expressed highly in brain and intestine, and moderately in kidney, gill, and eye of healthy olive flounder. Taken together, CaM may be necessary for early olive flounder development and that it may have a part in homeostasis.

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Changes in the levels of $Ca^{2+}$/calmodulin - binding proteins and glutamate decarboxylase during the growth of tobacco suspension cells (담배 배양 세포의 성장과정 중 칼슘/칼모듈린-결합단백질 및 glutamate decarboxylase의 생성변화)

  • Han, Kwang-Soo;Oh, Suk-Heung
    • Applied Biological Chemistry
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    • v.43 no.4
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    • pp.231-235
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    • 2000
  • The changes of calmodulin levels, calmodulin-binding proteins, and $Ca^{2+}$/calmodulin-dependent glutamate decarboxylase during the growth of tobacco suspension cells were investigated. Tobacco cells exhibited a typical growth curve, including an exponential growth phase between 3 and 5 days after inoculation, and an apparent stationary phase occurring after 5 day. Although slight changes were observed from sample to sample, calmodulin protein levels remained similar during the phases of culture growth. Several $Ca^{2+}-dependent$ calmodulin-binding proteins including 56, 46, 36, and 32-kDa proteins were detected in tobacco cell extracts. The 56-kDa protein was identified as glutamate decarboxylase by Western-blot analysis using an anti-GAD monoclonal antibody. The levels of GAD protein and the specific activity of GAD enzyme were highest during the middle exponential phase of the culture growth cycle. These data suggest that $Ca^{2+}$/calmodulin-dependent glutamate decarboxylase is modulated during the growth of tobacco suspension cells.

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Determination of $Ca^{2+}$ by Fiber Optic Fluorosensor Based on the Conformational Change of the Protein Calmodulin (Calmodulin 단백질의 형태변화를 이용한 광섬유 형광센서에 의한 $Ca^{2+}$의 정량)

  • Ri, Chang-Seop;Yang, Seung Tae
    • Analytical Science and Technology
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    • v.8 no.3
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    • pp.221-227
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    • 1995
  • The fiber optic fluorosensor that shows a specific selectivity for calcium ion is studied. This sensor employs protein Calmodulin(CaM) which forms a fluorescent chelate with $Ca^{2+}$. A dialysis membrane is used to entrap a fluorescein isothiocyanate-labeled CaM solution at the common end of a bifurcated fiber optic bundle. The sensing mechanism of this sensor is based on the shifts in the fluorescence spectrum of metal-calmodulin complexes which FCaM forms a chelate with $Ca^{2+}$. Upon binding with $Ca^{2+}$, CaM undergoes a conformational change which induces a change in the fluorescence of FCaM. This change in fluorescence signal which is measured by photomultiflier tube is related to the concentration of $Ca^{2+}$ for calibration curve. Detection limit for $Ca^{2+}$ and the interference effects by $Mg^{2+}$, $Eu^{3+}$ and $La^{3+}$ for this sensor are studied. Response time and life time for this fluorosensor are also investigated.

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Calcium Signaling-mediated and Differential Induction of Calmodulin Gene Expression by Stress in Oryza sativa L.

  • Phean-o-pas, Srivilai;Punteeranurak, Pornpimon;Buaboocha, Teerapong
    • BMB Reports
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    • v.38 no.4
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    • pp.432-439
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    • 2005
  • $Ca^{2+}$/calmodulin transduction pathways have been implicated in mediating stress response and tolerance in plants. Here, three genes encoding calmodulin (Cam) members of the EF-hand family of $Ca^{2+}$-binding proteins were identified from Oryza sativa L. databases. Complementary DNA for each of the calmodulin genes, OsCam1, OsCam2, and OsCam3 were sequenced. OsCam1 and OsCam2 encode a conventional 148-amino acid calmodulin protein that contains four characteristic $Ca^{2+}$-binding motifs. OsCam3 encode a similar protein with a 38-amino-acid extension containing a putative prenylation site (CVIL) at the carboxyl terminus. RT-PCR showed that each of the genes is expressed in leaves and roots of 2-week old rice seedlings. By RNA gel blot analysis, OsCam1 mRNA levels strongly increased in response to NaCl, mannitol and wounding treatments. In contrast, OsCam2 mRNA levels were relatively unchanged under all conditions investigated. NaCl treatment and wounding also increased the OsCam3 mRNA level, but in a more transient manner. Our results indicate that although the expression of genes encoding different calmodulin isoforms is ubiquitous, they are differentially regulated by various stress signals. In addition, we have demonstrated that the calcium-channel blocker lanthanum chloride inhibited the induction of OsCam1 gene expression by both NaCl and mannitol treatments. These results suggest that osmotic stress induced expression of OsCam1 gene requires the $[Ca^{2+}]_{cyt}$ elevation that is known to occur in response to these stimuli.

Prevention of Ischemic Damage in Working Rat Hearts by Calcium Channel Blocker and Calmodulin Inhibitors (흰쥐심장의 허혈손상에 대한 Calcium 통로봉쇄제와 Calmodulin 억제제의 예방효과에 대한 연구)

  • 성시찬
    • Journal of Chest Surgery
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    • v.22 no.6
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    • pp.901-913
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    • 1989
  • This study was investigated under the postulation that activation of intracellular calcium- calmodulin complex during ischemia-reperfusion leads to myocardial injury. The protective effects of calcium channel blocker, diltiazem and calmodulin inhibitors, trifluoperazine, flunarizine and calmidazolium from ischemic injury in rat hearts were observed by using Langendorff apparatus when the antagonists were infused for 3 min in the beginning of ischemia. Thereby, an increase in resting tension developed during 30-min ischemia was analyzed with regard to [1] the degree of cardiac functional recovery following 60-min reperfusion, [2] changes in biochemical variables evoked during 30-min ischemia. The results obtained were as follows: l. In the ischemic group, the resting tension was increased by 4.1*0.2 g at 30-min ischemia. However, the increase in resting tension was markedly reduced not only by pretreatment with diltiazem [3.3 p M] but also with calmodulin inhibitors, trifluoperazine [3.3 p M], flunarizine [0.5 p M] and calmidazolium [0.5 p M], respectively. 2. Recovery of myocardial contractility, +dF /dt and coronary flow were much reduced when evoked by reperfusion in the ischemic group. These variables were significantly improved either by pretreatment with diltiazem or with calmodulin inhibitors. 3. The resting tension increment evoked during ischemia was significantly inversely correlated with the degree of cardiac function recovered during reperfusion. 4. Following 30-min ischemia, the production of malondialdehyde and release of lysosomal enzyme were much increased in association with a decrease in creatine kinase activity. 5. The increases in malondialdehyde production and release of free lysosomal enzyme were suppressed by pretreatment with calmodulin inhibitors as well as diltiazem. Likewise, the decrease of creatine kinase activities was prevented by these calcium antagonists. With these results, it is indicated that a increase in resting tension observed during ischemia has an inverse relationship to the cardiac function recovered following reperfusion, and further, the later may be significantly dependent on the degree of biochemical alterations occurred during ischemia such as decrease in creatine kinase activity, increased production of malondialdehyde and increased release of free lysosomal enzyme. Thus it is concluded that calmodulin plays a pivotal role in the process of ischemic injury.

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Isolation and Characterization of Calmodulin 2 (CICAM2) Gene from Codonopsis lanceolata

  • Lee, Kang;In, Jun-Gyo;Yu, Chang-Yeon;Min, Byung-Hoon;Chung, Ill-Min;Kim, Se-Young;Kim, Yeong-Chae;Yang, Deok-Chun
    • Plant Resources
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    • v.7 no.3
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    • pp.174-180
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    • 2004
  • Calmodulin, a $Ca^{2+}$-binding protein, has no enzyme activity. It combines with $Ca^{2+}$ and makes variable proteins to an active form. Calmodulin 2 is a ubiquitous protein in plants. To investigate the defense mechanism against various stresses, a clone encoding a calmodulin 2 protein was isolated from a cDNA library prepared from taproot mRNAs of Codonopsis lanceolata. The cDNA, designated CICAM2, is 719 nucleotides long and has an open reading frame of 450 bp with a deduced amino acid sequence of 149 residues. The deduced amino acid sequence of CICAM2 showed a high similarity with calmodulins of P. x hybrida (P27163) 97%, N. tabacum (BAB61908) 97%, S. tuberosum (AAA74405) 96%, Z. mays (CAA74307) 92%, C. richardii (AF510075) 93%, M. truncatula (AAM81203) 91%, and G. max (P62163) 91%. The transcriptional expression of the CICAM2 gene, was gradually increased by the CaCl$_2$ treatment. Whereas its expression And it was gradually decreased in the cold stress treatment.ent.

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Tissue Specific Expression of Wound-Inducible RCaM-2 Promoter in Transgenic Tobacco Plants (상처에 의해서 유도되는 벼 calmodulin promoter의 transgenic 담배에서조직 특이적 발현)

  • Choi Young Ju
    • Journal of Life Science
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    • v.15 no.2 s.69
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    • pp.176-181
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    • 2005
  • To study calmodulin (CaM) gene expression and its regulation, rice CaM promoter (RCaM-2) was isolated and fused to $\beta-glucuronidase$ (GUS), reporter gene. X-Glue staining patterns revealed that GUS localization is high in meristemic tissues such as the stem apex, stolen tip, and vascular regions. GUS staining in the transverse sections of stem and petiole was restricted to the inside of the vascular system, and cortex and epidermis located outside of the vascular system usually did not show GUS staining even a plant that expressed strong activity. GUS activity was found to be tissue specific expressed and exhibited a dramatic transient increase in response to wounding. These results suggest that the 5'-flanking region of RCaM gene regulates wound-inducible expression.

Isolation and Characterization of a Novel Calcium/Calmodulin-Dependent Protein Kinase, AtCK, from Arabidopsis

  • Jeong, Jae Cheol;Shin, Dongjin;Lee, Jiyoung;Kang, Chang Ho;Baek, Dongwon;Cho, Moo Je;Kim, Min Chul;Yun, Dae-Jin
    • Molecules and Cells
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    • v.24 no.2
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    • pp.276-282
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    • 2007
  • Protein phosphorylation is one of the major mechanisms by which eukaryotic cells transduce extracellular signals into intracellular responses. Calcium/calmodulin ($Ca^{2+}/CaM$)-dependent protein phosphorylation has been implicated in various cellular processes, yet little is known about $Ca^{2+}/CaM$-dependent protein kinases (CaMKs) in plants. From an Arabidopsis expression library screen using a horseradish peroxidase-conjugated soybean calmodulin isoform (SCaM-1) as a probe, we isolated a full-length cDNA clone that encodes AtCK (Arabidopsis thaliana calcium/calmodulin-dependent protein kinase). The predicted structure of AtCK contains a serine/threonine protein kinase catalytic domain followed by a putative calmodulin-binding domain and a putative $Ca^{2+}$-binding domain. Recombinant AtCK was expressed in E. coli and bound to calmodulin in a $Ca^{2+}$-dependent manner. The ability of CaM to bind to AtCK was confirmed by gel mobility shift and competition assays. AtCK exhibited its highest levels of autophosphorylation in the presence of 3 mM $Mn^{2+}$. The phosphorylation of myelin basic protein (MBP) by AtCK was enhanced when AtCK was under the control of calcium-bound CaM, as previously observed for other $Ca^{2+}/CaM$-dependent protein kinases. In contrast to maize and tobacco CCaMKs (calcium and $Ca^{2+}/CaM$-dependent protein kinase), increasing the concentration of calmodulin to more than $3{\mu}M$ suppressed the phosphorylation activity of AtCK. Taken together our results indicate that AtCK is a novel Arabidopsis $Ca^{2+}/CaM$-dependent protein kinase which is presumably involved in CaM-mediated signaling.

Depression of $Ca^{2+}$ Influx in Complement C5a-stimulated Neutrophils by Calmodulin Inhibitors

  • Ham, Dong-Suk;Kim, Hyun-Ho;Han, Eun-Sook;Lee, Chung-Soo
    • The Korean Journal of Physiology and Pharmacology
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    • v.2 no.1
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    • pp.109-117
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    • 1998
  • Role of $Ca^{2+}$/calmodulin complex in intracellular $Ca^{2+}$ mobilization in neutrophils has not been clearly elucidated. In this study, effects of chlorpromazine, trifluoperazine and imipramine on the intracellular $Ca^{2+}$ mobilization, including $Ca^{2+}$ influx, in C5a-activated neutrophils were investigated. Complement C5a- stimulated superoxide production and myeloperoxidase release in neutrophils were inhibited by chlorpromazine, trifluoperazine and imipramine, except no effect of imipramine on myeloperoxidase release. A C5a-elicited elevation of [$Ca^{2+}$]i in neutrophils was inhibited by chlopromazine, trifluoperazine, imipramine, staurosporine, genistein, EGTA, and verapamil but not affected by pertussis toxin. The intracellular $Ca^{2+}$ release in C5a-activated neutrophils was not affected by chlorpromazine and imipramine. Chlorpromazine and imipramine inhibited $Mn^{2+}$ influx by C5a-activated neutrophils. Thapsigargin-evoked $Ca^{2+}$ entry was inhibited by chlorpromazine, trifluoperazine, imipramine, genistein, EGTA and verapamil, while the effect of staurosporine was not detected. The results suggest that $Ca^{2+}$/calmodulin complex is involved in the activation process of neutrophils. The depressive action of calmodulin inhibitors on the elevation of cytosolic $Ca^{2+}$ level in C5a-activated neutrophils appears to be accomplished by inhibition of $Ca^{2+}$ influx from the extracellular medium.

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