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Activation of the cardiac Na(+)-Ca(2+) exchanger by sorcin via the interaction of the respective Ca(2+)-binding domains.

Zamparelli C, Macquaide N, Colotti G, Verzili D, Seidler T, Smith GL, Chiancone E - J. Mol. Cell. Cardiol. (2010)

Bottom Line: To investigate the importance of this region in the interaction with NCX1, three variants were examined: W105G and W99G, mutated respectively near EF3 and EF2, and E124A that does not bind Ca(2+) due to a mutation at EF3.Downregulation of sorcin decreased and supplementation with wt sorcin (3muM) increased NCX activity in isolated cardiomyocytes.The relative stimulatory effects of the sorcin variants were: W105G>wt sorcin>Sorcin Calcium Binding Domain (SCBD)>W99G>E124A.

View Article: PubMed Central - PubMed

Affiliation: C.N.R. Institute of Molecular Biology and Pathology, Department of Biochemical Sciences A. Rossi Fanelli, Sapienza University of Rome, 00185 Rome, Italy. emilia.chiancone@uniroma1.it

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Binding of wild type sorcin and sorcin variants to immobilized CBD1. Concentration of wt sorcin and variants was 6 μM, in 10 mM HEPES, 0.15 M NaCl, 50 μM CaCl2, and 0.005% surfactant P-20, at pH 7.4. The increase in RU relative to baseline indicates complex formation, whereas the decrease in RU represents dissociation of sorcin from the immobilized CBD1 domain after injection of buffer. The temperature was 25 °C. The sensorgrams shown are the average of two experiments. From top to bottom: W105G, SCBD, wt sorcin, W99G, and E124A.
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fig8: Binding of wild type sorcin and sorcin variants to immobilized CBD1. Concentration of wt sorcin and variants was 6 μM, in 10 mM HEPES, 0.15 M NaCl, 50 μM CaCl2, and 0.005% surfactant P-20, at pH 7.4. The increase in RU relative to baseline indicates complex formation, whereas the decrease in RU represents dissociation of sorcin from the immobilized CBD1 domain after injection of buffer. The temperature was 25 °C. The sensorgrams shown are the average of two experiments. From top to bottom: W105G, SCBD, wt sorcin, W99G, and E124A.

Mentions: The NCX activity data just presented were supplemented by SPR experiments in which the interaction between immobilized CBD1 and soluble wt sorcin or sorcin variants at the same protein concentration was monitored (Fig. 8). In the presence of 50 µM Ca2+, the interaction with CBD1 follows the order W105G > SCBD > wt sorcin > W99G based on the observed increase in RU. The corresponding KD values decrease from 26 to 2.0 μM (average of two experiments). In case of the E124A variant the increase in RU was negligible.


Activation of the cardiac Na(+)-Ca(2+) exchanger by sorcin via the interaction of the respective Ca(2+)-binding domains.

Zamparelli C, Macquaide N, Colotti G, Verzili D, Seidler T, Smith GL, Chiancone E - J. Mol. Cell. Cardiol. (2010)

Binding of wild type sorcin and sorcin variants to immobilized CBD1. Concentration of wt sorcin and variants was 6 μM, in 10 mM HEPES, 0.15 M NaCl, 50 μM CaCl2, and 0.005% surfactant P-20, at pH 7.4. The increase in RU relative to baseline indicates complex formation, whereas the decrease in RU represents dissociation of sorcin from the immobilized CBD1 domain after injection of buffer. The temperature was 25 °C. The sensorgrams shown are the average of two experiments. From top to bottom: W105G, SCBD, wt sorcin, W99G, and E124A.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2958309&req=5

fig8: Binding of wild type sorcin and sorcin variants to immobilized CBD1. Concentration of wt sorcin and variants was 6 μM, in 10 mM HEPES, 0.15 M NaCl, 50 μM CaCl2, and 0.005% surfactant P-20, at pH 7.4. The increase in RU relative to baseline indicates complex formation, whereas the decrease in RU represents dissociation of sorcin from the immobilized CBD1 domain after injection of buffer. The temperature was 25 °C. The sensorgrams shown are the average of two experiments. From top to bottom: W105G, SCBD, wt sorcin, W99G, and E124A.
Mentions: The NCX activity data just presented were supplemented by SPR experiments in which the interaction between immobilized CBD1 and soluble wt sorcin or sorcin variants at the same protein concentration was monitored (Fig. 8). In the presence of 50 µM Ca2+, the interaction with CBD1 follows the order W105G > SCBD > wt sorcin > W99G based on the observed increase in RU. The corresponding KD values decrease from 26 to 2.0 μM (average of two experiments). In case of the E124A variant the increase in RU was negligible.

Bottom Line: To investigate the importance of this region in the interaction with NCX1, three variants were examined: W105G and W99G, mutated respectively near EF3 and EF2, and E124A that does not bind Ca(2+) due to a mutation at EF3.Downregulation of sorcin decreased and supplementation with wt sorcin (3muM) increased NCX activity in isolated cardiomyocytes.The relative stimulatory effects of the sorcin variants were: W105G>wt sorcin>Sorcin Calcium Binding Domain (SCBD)>W99G>E124A.

View Article: PubMed Central - PubMed

Affiliation: C.N.R. Institute of Molecular Biology and Pathology, Department of Biochemical Sciences A. Rossi Fanelli, Sapienza University of Rome, 00185 Rome, Italy. emilia.chiancone@uniroma1.it

Show MeSH
Related in: MedlinePlus