scholarly journals Mechanism of inhibition of the calcium pump of sarcoplasmic reticulum by thapsigargin

1992 ◽  
Vol 283 (2) ◽  
pp. 525-529 ◽  
Author(s):  
M Wictome ◽  
I Henderson ◽  
A G Lee ◽  
J M East
Keyword(s):  
Steady State ◽  
Sarcoplasmic Reticulum ◽  
Atpase Activity ◽  
Dissociation Constant ◽  
State Level ◽  
Calcium Pump ◽  
Molar Ratio ◽  
Steady State Level ◽  
Mechanism Of Inhibition ◽  
Nanomolar Range

The steady-state ATPase activity of sarcoplasmic-reticulum (Ca(2+)-Mg2+)-ATPase is inhibited by thapsigargin at a molar ratio of 1:1, with a dissociation constant for thapsigargin estimated to be in the sub-nanomolar range. In the presence of thapsigargin, only a single Ca2+ ion binds to the ATPase. Similarly, addition of thapsigargin to the ATPase incubated in the presence of Ca2+ results in the release of one of the two originally bound Ca2+ ions. As monitored by the fluorescence of nitrobenzo-2-oxa-1,3-diazole-labelled ATPase, thapsigargin appears to shift the transition between E1 and E2 conformations towards E2. Addition of thapsigargin prevents phosphorylation of the ATPase by P(i) and results in a very low steady-state level of phosphorylation of the ATPase by ATP, as observed previously for nonylphenol.

scholarly journals Effects of Ca2+, Mg2+ and calmodulin on the formation and decomposition of the phosphorylated intermediate of the erythrocyte Ca2+-stimulated ATPase

1987 ◽  
Vol 244 (3) ◽  
pp. 617-623 ◽  
Author(s):  
B G Allen ◽  
S Katz ◽  
B D Roufogalis
Keyword(s):  
Acetic Acid ◽  
Steady State ◽  
Atpase Activity ◽  
Human Erythrocyte ◽  
Lower Energy ◽  
State Level ◽  
Steady State Level

Formation of the phosphorylated intermediate (ECaP) of the human erythrocyte Ca2+-stimulated ATPase (Ca2+-ATPase) was more rapid and reached steady state sooner at 400 microM-Ca2+ than at 1 microM-Ca2+. Calmodulin increased the apparent rate of ECaP formation at 1 microM-Ca2+, whereas at 400 microM-Ca2+, calmodulin decreased the steady-state level of the ECaP without affecting its apparent rate of formation. Removal of endogenous Mg2+ with trans-1,2-diaminocyclohexane-NNN'N'-tetra-acetic acid, which decreased both the velocity and Ca2+-sensitivity of the Ca2+-ATPase, did not alter the Ca2+-sensitivity or the apparent rate of formation of ECaP. ECaP formation at high Ca2+ concentrations was not affected by Mg2+ concentrations as high as 1 mM, and the ECaP could be dephosphorylated by ADP and ATP along either the forward or reverse pathways. The results suggest that high Ca2+ concentrations inhibit Ca2+-ATPase activity by preventing dephosphorylation of the E2P complex, rather than by inhibition of the transformation from E1CaP (‘high-Ca2+-affinity’ ECaP) to E2CaP (‘lower-energy’ ECaP).

Tacrolimus initial steady state level in post-transplant cyclophosphamide-based GvHD prophylaxis regimens

2021 ◽  
Author(s):  
Janny M. Yao ◽  
Dongyun Yang ◽  
Mary C. Clark ◽  
Salman Otoukesh ◽  
Thai Cao ◽  
...  
Keyword(s):  
Steady State ◽  
State Level ◽  
Steady State Level ◽  
Post Transplant ◽  
Gvhd Prophylaxis

Ascorbate and glutathione homeostasis in vascular smooth muscle cells: cooperation with endothelial cells

1998 ◽  
Vol 275 (4) ◽  
pp. C1031-C1039 ◽  
Author(s):  
Ilia Voskoboinik ◽  
Karin Söderholm ◽  
Ian A. Cotgreave
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Steady State ◽  
Smooth Muscle Cells ◽  
Umbilical Vein ◽  
State Level ◽  
Muscle Cells ◽  
Free Access ◽  
Steady State Level ◽  
Human Umbilical Vein

Human umbilical vein smooth muscle cells (HUVSMCs) utilize extracellular cystine, glutathione (GSH), and N-acetylcysteine (NAC) to synthesize cellular GSH. Extracellular cystine was effective from 5 μM, whereas GSH and NAC were required at 100 μM for comparable effects. The efficacy of extracellular GSH was dependent on de novo GSH synthesis, indicating a dependence on cellular γ-glutamyltransferase (glutamyl transpeptidase). Coculture of syngenetic HUVSMCs and corresponding human umbilical vein endothelial cells (HUVECs) on porous supports restricted cystine- or GSH-stimulated synthesis of HUVSMC GSH when supplied on the “luminal” endothelial side. Thus HUVSMC GSH rapidly attained a steady-state level below that achieved in the absence of interposed HUVECs. HUVSMCs also readily utilize both reduced ascorbate (AA) and oxidized dehydroascorbate (DHAA) over the range 50–500 μM. Phloretin effectively blocked both AA- and DHAA-stimulated assimilation of intracellular AA, indicating a role for a glucose transporter in their transport. Uptake of extracellular AA was also sensitive to extracellular, but not intracellular, thiol depletion. When AA was applied to the endothelial side of the coculture model, assimilation of intracellular AA in HUVSMCs was restricted to a steady-state level below that achieved by free access.

scholarly journals The steady-state level of Mg-protoporphyrin IX is not a determinant of plastid-to-nucleus signaling in Arabidopsis

2008 ◽  
Vol 105 (39) ◽  
pp. 15184-15189 ◽  
Author(s):  
N. Mochizuki ◽  
R. Tanaka ◽  
A. Tanaka ◽  
T. Masuda ◽  
A. Nagatani
Keyword(s):  
Steady State ◽  
Protoporphyrin Ix ◽  
State Level ◽  
Steady State Level

scholarly journals Expression of human beta-secretase in the mouse brain increases the steady-state level of beta-amyloid

2002 ◽  
Vol 80 (5) ◽  
pp. 799-806 ◽  
Author(s):  
Ursula Bodendorf ◽  
Simone Danner ◽  
Frauke Fischer ◽  
Muriel Stefani ◽  
Christine Sturchler-Pierrat ◽  
...  
Keyword(s):  
Steady State ◽  
Mouse Brain ◽  
State Level ◽  
Beta Amyloid ◽  
Steady State Level ◽  
Beta Secretase

Expression of a gene for mouse eucaryotic elongation factor Tu during murine erythroleukemic cell differentiation

1987 ◽  
Vol 7 (11) ◽  
pp. 3929-3936
Author(s):  
W W Roth ◽  
P W Bragg ◽  
M V Corrias ◽  
N S Reddy ◽  
J N Dholakia ◽  
...  
Keyword(s):  
Steady State ◽  
Genomic Library ◽  
Elongation Factor ◽  
State Level ◽  
Genomic Clone ◽  
Elongation Factor Tu ◽  
Steady State Level ◽  
Similar Reduction ◽  
S1 Nuclease ◽  
Murine Erythroleukemia

The eucaryotic elongation factor Tu (eEF-Tu) is a single polypeptide with an approximate Mr of 53,000. During protein synthesis eEF-Tu promotes the binding of aminoacyl-tRNA to the ribosome. To study the expression of the gene(s) for this factor, a genomic clone was isolated that contains a mouse eEF-Tu gene. We screened a phage genomic library with a synthetic oligonucleotide probe complementary to a region of the Saccharomyces cerevisiae and Artemia sp. eEF-Tu genes which codes for an area that is highly conserved between both yeast and Artemia sp. eEF-Tu. From approximately 75,000 phage plaques we obtained five isolates with apparently identical inserts. All five clones contained a 3.8-kilobase EcoRI fragment that hybridized to additional oligonucleotide probes corresponding to different conserved regions of eEF-Tu. We sequenced the 5' end of one genomic clone and determined the length of the cloned fragment that was protected by eEF-Tu mRNA in S1 nuclease protection assays. A quantitative S1 nuclease protection assay was used to compare the relative steady-state levels of eEF-Tu mRNA in total mRNA in total RNA isolated from hexamethylene-bisacetamide-induced murine erythroleukemia cells. The results show a dramatic reduction in the steady-state level of eEF-Tu mRNA as differentiation proceeds. A similar reduction in transcription of eEF-Tu mRNA was observed in isolated nuclei. Finally, we examined the in vivo synthesis of eEF-Tu during differentiation and found that it declined in a manner parallel to the decline in the steady-state level of eEF-Tu mRNA. In addition, we have isolated and sequenced a cDNA clone for mouse eEF-Tu. The derived amino acid sequence is compared with sequences from other eucaryotes.

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