• Title/Summary/Keyword: calmodulin 2

Search Result 196, Processing Time 0.022 seconds

Changes in Kinetic Properties of $Ca^{2+}$/Calmodulin-Dependent Protein Kinase la Activated by $Ca^{2+}$/Calmodulin-Dependent Protein Kinase I Kinase (칼슘/칼모듈린-의존성 단백질 키나아제 I 키나아제에 의한 칼슘/칼모듈린-의존성 단백질 키나아제 Ia의 활성화에 따른 효소반응 특성의 변화)

  • Cho, Jung-Sook
    • YAKHAK HOEJI
    • /
    • v.41 no.6
    • /
    • pp.773-781
    • /
    • 1997
  • The activity of $Ca^{2+}$calmodulin (CaM)-dependent protein kinase Ia (CaM kinase Ia) is shown to be regulated through direct phosphorylation by CaM kinase I kinase (CaMK IK). In the present study, three distinct CaMKIK peaks were separated from Q-Sepharose colunm chromatography of pig brain homogenate using a Waters 650 Protein Purification System. The purified CaMKIK from the major peak potently and rapidly enhanced CaM kinase Ia activity, reaching a maximal stimulation within 2min at the concentrations of 12-15nM. The activated state of CaM kinase Ia is characterized by a markedly enhanced $V_{max}4 as well as significantly decreased $K_m\;and\;K_a$ values toward peptide substrate and CaM, respectively. These observations suggest the activation process of CaM kinase Ia. The phosphorylation of CaM kinase Ia by CaMKIK may induce its conformational change responsible for the alterations in the kinetic properties, which ultimately leads to the rapid enzyme activation.

  • PDF

The Role of $Ca^{2+}$/Calmodulin-Dependent Protein Kinase II on the Norepinephrine and GTP-Increased Myosin tight Chain Phosphorylations in Rabbit Mesenteric ${\alpha}-toxin$ Permeabilized Artery (${\alpha}$-독으로 처리한 토끼창간막동맥에서 Norepinephrine과 GTP에 의한 마이오신 인산화의 증가에 대한 $Ca^{2+}$/calmodulin-dependent Protein Kinase II의 역할)

  • Ahn, Hee-Yul;Kim, Hun-Sik;Moreland, Robert S.
    • The Korean Journal of Pharmacology
    • /
    • v.30 no.1
    • /
    • pp.111-116
    • /
    • 1994
  • The role of $Ca^{2+}$/calmodulin-dependent protein kinase II in the increase of myofilament $Ca^{2+}$ sensitivity by agonist and GTP was investigated in rabbit mesenteric ${\alpha}-toxin$ permeabilized artery. $0.3{\mu}M\;Ca^{2+}$ increased myosin light chain phosphorylations monotonically. $10\;{\mu}M$ norepinephrine and $10\;{\mu}M$ GTP potentiated increase of myosin light chain phosphorylations by $0.3{\mu}M\;Ca^{2+}$, which reaches a peak at 5 min and gradually declines to the $Ca^{2+}$ alone level at 20 min. At the early phase (1 min), $10\;{\mu}M$ KN 62, the inhibitor of $Ca^{2+}$/calmodulin-dependent protein kinase II , decreased myosin light chain phosphorylation levels by $10\;{\mu}M$ norepinephrine and $10\;{\mu}M$ GTP in the presence of $0.3{\mu}M\;Ca^{2+}.\;However\;10\;{\mu}M$ KN-62 did not affect the myosin light chain phosphorylations by $10\;{\mu}M$ norepinephrine and $10\;{\mu}M$ GTP in the presence of $0.3{\mu}M\;Ca^{2+}$ at the peak (5 min) and plateau phases (20 min). From these results, the role of $Ca^{2+}$/calmodulin-dependent protein kinase II may be different depending on time, which may play a role in increase of myofilamint $Ca^{2+}$ sensitivity by norepinephrine and GTP resulting from increase of myosin light chain phosphorylations at the early phase. However, at plateau phase, $Ca^{2+}$/calmodulin-dependent protein kinase II may not be involved in the increase of myofilament $Ca^{2+}$ sensitivity by norepinephrine and GTP in rabbit mesenteric ${\alpha}-toxin$ permeabilized artery.

  • PDF

Relation of $\Ca^{2+}$-ATPase and trigger peptidase(TPase) that are Membrane Proteins in a Differentiation Process on Heterobasidiomycerous Yeast (이담자 효모균의 성분화과정에서 막단백질 중 $\Ca^{2+}$-ATPase와 trigger peptidase(TPase)의 상호관계)

  • 정영기;이태호;정경태
    • Microbiology and Biotechnology Letters
    • /
    • v.22 no.1
    • /
    • pp.1-6
    • /
    • 1994
  • We have studied the relation between Ca$^{2+}$-ATPase and trigger peptidase(TPase) which are membeane protein well known as their significant role for signal transduction of mating pheromone in heterobasidiomycetous yeast. Rhodosporidium toruloides. We found out that there were Ca $^{2+}$-ATPase and TPase together in isolated calmodulim binding protein(CBP), usion calmodulin affinity column chromatography after solubilization of mation type a cell membrane protein, and that the dependence of enzyme activity of both the enzymes on Ca$^{2+}$, phospholipid and nonionic detergent are similar. However, Ca$^{2+}$-ATPase hed quite absolute dependence on calmodulin and, on the other hand, TPase didn't have any dependence. Judging from the fact that there are both enzymes in CBP which the dependence of calmodulin are quite different, we found out that both enzymes were made to their compound and existed in mating type a cell membrane.

  • PDF

Molecular Characterization of Three cDNA Clones Encoding Calmodulin Isoforms of Rice

  • Lee, Sung-Ho;Kim, Cha Young;Lim, Chae Oh;Lee, Soo In;Gal, Sang Wan;Choi, Young Ju
    • Journal of Applied Biological Chemistry
    • /
    • v.43 no.1
    • /
    • pp.5-11
    • /
    • 2000
  • Three cDNA clones encoding rice calmodulin (CaM) isoforms (OsCaM-1, OsCaM-2, and OsCaM-3) were isolated from a rice cDNA library constructed from suspension-cultured rice cells treated with fungal elicitor. The coding regions of OsCaM-1 and O.sCaM-2 were 89% homologous at DNA Ievel, whereas the 5' and 3' untranslated regions were highly divergent. The polypeptides encoded by OsCaM-1 and OsCaM-2 was identical except two conservative substitution at position 8 and 75. The coding region of OsCaM-3 was consist of a typical conserved CaM domain and an additional C-terminal extension. The amino acid sequence of conserved CaM domain of OsCaM-3 shared only 86% identity with that OsCaM-1. The OsCaM-3 cDNA is belongs to a novel group of calmodulin gene due to its C-terminal extension of 38 amino acids, a large number of which are positively charged. The extension also contains a C-terminal CaaX-box prenylation site (CVlL). Genomic Southern analysis revealed at least six copies of CaM or CaM-related genes, suggesting that calmodulin may be represented by a small multigene family in the rice geneme. Expression of OsCaM gene was examined through Northern blot analysis. Transcript level of OsCaM-3 was increased by treatment with a fungal elicitor, whereas the OsCaM-1 and OsCaM-2 genes did not respond to the fungal elicitor. The expression of OsCaM-3 gene was remarkable inhibited in the rice cells treated with cyclosporine A, calcinurin inhibitor.

  • PDF

Structure and expression analysis of the OsCam1-1 calmodulin gene from Oryza sativa L.

  • Phean-o-pas, Srivilai;Limpaseni, Tipaporn;Buaboocha, Teerapong
    • BMB Reports
    • /
    • v.41 no.11
    • /
    • pp.771-777
    • /
    • 2008
  • Calmodulin (CaM) proteins, members of the EF-hand family of $Ca^{2+}$-binding proteins, represent important relays in plant calcium signals. Here, OsCam1-1 was isolated by PCR amplification from the rice genome. The gene contains an ORF of 450 base pairs with a single intron at the same position found in other plant Cam genes. A promoter region with a TATA box at position-26 was predicted and fused to a gus reporter gene, and this construct was used to produce transgenic rice by Agrobacterium-mediated transformation. GUS activity was observed in all organs examined and throughout tissues in cross-sections, but activity was strongest in the vascular bundles of leaves and the vascular cylinders of roots. To examine the properties of OsCaM1-1, the encoding cDNA was expressed in Escherichia coli. The electrophoretic mobility shift when incubated with $Ca^{2+}$ indicates that recombinant OsCaM1-1 is a functional $Ca^{2+}$-binding protein. In addition, OsCaM1-1 bound the CaMKII target peptide confirming its likely functionality as a calmodulin.

Environmental effects on plant calmodulin system (식물 칼모듈린 체계에 미치는 환경적 요인의 영향)

  • Yang, Moon-Sik;Oh, Suk-Heung
    • Applied Biological Chemistry
    • /
    • v.39 no.1
    • /
    • pp.25-31
    • /
    • 1996
  • Transgenic tobacco plants expressing calmodulin derivative($lys{\rightarrow}ile$ 115 calmodulin) and hygromycin resistance genes or hygromycin resistance gene alone(control) were generated by Agrobacterium-mediated DNA transfer. Seeds obtained from the transgenic plants($F_o$) were tested for resistance to hygromycin and the expected 3 : 1 ratio was observed. The expression of calmodulin derivative in the tobacco plants was identified by a combined method of Western blot and Chemiluminescence. The effects of surface sterilizers on the germiation of seeds from transgenic tobacco plants were tested in Murashige and Skoog agar medium. Seeds obtained from transgenic tobacoo plants expressing the calmodulin derivative showed no fungi contamination with normal germination by treating with sterilized water alone or sodium hypochlorite(2% effective chlorine). In contrast, seeds from the control transgenic tobacco plants showed severe contamination with fungi by treating with sterilized water alone and showed no contamination with normal germination by treating with sodium hypochlorite(2% chlorine). The effects of calcium concentration on the germination of seeds from transgenic tobacco plants were tested in Murashige and Skoog agar medium. Seeds obtained from transgenic tobacco plants expressing the calmodulin derivative showed better germination frequency than that of the control transgenic tobacco seeds in the medium containing 30 mM $CaCl_2$. The data raise the possibility that the expression of calmodulin derivative gene in tobacco plants could increase adaptability of the seeds to environmental stresses.

  • PDF

Determination of Eu(III) by Fluorescence Spectrometry using Fiber Optic Sensor (광섬유센서를 이용한 Eu(III)의 형광분광법적 정량)

  • Lee, Sang Hak;Lee, Yoon Hee;Yang, Seung Tae;Choi, Sang Seob
    • Analytical Science and Technology
    • /
    • v.11 no.5
    • /
    • pp.409-412
    • /
    • 1998
  • The analytical method to determine europium(III) ion in aqueous solution by fluorescence spectroscopy based upon the conformational change of calmodulin in the presence of the analyte has been studied. The fiber optic chemical sensor used in this study was constructed by entrapping a fluorescein-labeled calmodulin solution, EGTA, buffer solution at the common end of a bifurcated fiber optic bundle by means of a dialysis membrane. The calibration curve to determine europium(III) ion was obtained when concentration of calmodulin, concentration of EGTA, Tris-HCl buffer solution, pH, excitation wavelength and fluorescence wavelength were $5.0{\times}10^{-5}M$, 0.50 mM, 5.0 mM, 7.0, 495 nm and 520 nm, respectively. The detection limit was $1.0{\times}10^{-11}M$ and the working range of the calibration curve for the sensor was $1.0{\times}10^{-11}M{\sim}1.0{\times}10^{-9}M$. The response time was 15 minutes. For the determination of europium(III) ion by the present method, $Na^+$ and $K^+$ ions did not interfere but $Ca^{2+}$ ion seriously interfered.

  • PDF

Ca2+/calmodulin-dependent regulation of polycystic kidney disease 2-like-1 by binding at C-terminal domain

  • Baik, Julia Young;Park, Eunice Yon June;So, Insuk
    • The Korean Journal of Physiology and Pharmacology
    • /
    • v.24 no.3
    • /
    • pp.277-286
    • /
    • 2020
  • Polycystic kidney disease 2-like-1 (PKD2L1), also known as polycystin-L or TRPP3, is a non-selective cation channel that regulates intracellular calcium concentration. Calmodulin (CaM) is a calcium binding protein, consisting of N-lobe and C-lobe with two calcium binding EF-hands in each lobe. In previous study, we confirmed that CaM is associated with desensitization of PKD2L1 and that CaM N-lobe and PKD2L1 EF-hand specifically are involved. However, the CaM-binding domain (CaMBD) and its inhibitory mechanism of PKD2L1 have not been identified. In order to identify CaM-binding anchor residue of PKD2L1, single mutants of putative CaMBD and EF-hand deletion mutants were generated. The current changes of the mutants were recorded with whole-cell patch clamp. The calmidazolium (CMZ), a calmodulin inhibitor, was used under different concentrations of intracellular. Among the mutants that showed similar or higher basal currents with that of the PKD2L1 wild type, L593A showed little change in current induced by CMZ. Co-expression of L593A with CaM attenuated the inhibitory effect of PKD2L1 by CaM. In the previous study it was inferred that CaM C-lobe inhibits channels by binding to PKD2L1 at 16 nM calcium concentration and CaM N-lobe at 100 nM. Based on the results at 16 nM calcium concentration condition, this study suggests that CaM C-lobe binds to Leu-593, which can be a CaM C-lobe anchor residue, to regulate channel activity. Taken together, our results provide a model for the regulation of PKD2L1 channel activity by CaM.

Role of $Ca^{2+}$ and Calmodulin on the Initiation of Sperm Motility in Salmonid Fishes

  • Kho, Kang-Hee;Morisawa, Masaaki;Choi, Kap-Seong
    • Journal of Microbiology and Biotechnology
    • /
    • v.14 no.3
    • /
    • pp.456-465
    • /
    • 2004
  • $K^+$ efflux through a certain type of $K^+$ channels causes the change of membrane potential and leads to cAMP synthesis in the transmembrane cell signaling for the initiation of sperm motility in the salmonid fishes. The addition of $Ca^{2+}$ conferred motility to the trout sperm that were immobilized by external $K^+$ and other alkaline metals, $Rb^+$ and $Cs^{2+}$, suggesting the participation of external $Ca^{2+}$ in the initiation of sperm motility. L-type $Ca^{2+}$ channel blockers such as nifedipine, nimodipine, and FS-2 inhibited the motility, but N-type $Ca^{2+}$ channel blocker, w-conotoxin MvIIA, did not. On the other hand, the membrane hyperpolarization and cAMP synthesis were suppressed by $Ca^{2+}$ channel blockers, nifedipine, and trifluoroperazine. Furthermore, these suppressions were relieved by the addition of $K^+$ ionophore, valinomycin. Inhibitors of calmodulin, such as W-7, trifluoperazine, and calrnidazol-C1, inhibited the sperm motility, membrane hyperpolarization, and cAMP synthesis. The results suggest that $Ca^{2+}$ influx through $Ca^{2+}$ channels that are sensitive to specific $Ca^{2+}$ channel blockers and calmodulin participate in the changes of membrane potential, leading to synthesis of cAMP in the cell signaling for the initiation of trout sperm motility.

Increased calcium-mediated cerebral processes after peripheral injury: possible role of the brain in complex regional pain syndrome

  • Nahm, Francis Sahngun;Lee, Jae-Sung;Lee, Pyung-Bok;Choi, Eunjoo;Han, Woong Ki;Nahm, Sang-Soep
    • The Korean Journal of Pain
    • /
    • v.33 no.2
    • /
    • pp.131-137
    • /
    • 2020
  • Background: Among various diseases that accompany pain, complex regional pain syndrome (CRPS) is one of the most frustrating for patients and physicians. Recently, many studies have shown functional and anatomical abnormalities in the brains of patients with CRPS. The calcium-related signaling pathway is important in various physiologic processes via calmodulin (CaM) and calcium-calmodulin kinase 2 (CaMK2). To investigate the cerebral mechanism of CRPS, we measured changes in CaM and CaMK2 expression in the cerebrum in CRPS animal models. Methods: The chronic post-ischemia pain model was employed for CRPS model generation. After generation of the animal models, the animals were categorized into three groups based on changes in the withdrawal threshold for the affected limb: CRPS-positive (P), CRPS-negative (N), and control (C) groups. Western blot analysis was performed to measure CaM and CaMK2 expression in the rat cerebrum. Results: Animals with a decreased withdrawal threshold (group P) showed a significant increment in cerebral CaM and CaMK2 expression (P = 0.013 and P = 0.021, respectively). However, groups N and C showed no difference in CaM and CaMK2 expression. Conclusions: The calcium-mediated cerebral process occurs after peripheral injury in CRPS, and there can be a relationship between the cerebrum and the pathogenesis of CRPS.