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• W2162078918 abstract "The Na(+)-Ca(2+) exchanger in mammalian heart muscle (NCX1) is the central transporter protein that regulates extrusion of Ca(2+) from the heart cell. However, the functional biochemistry and physiology of NCX1 have been severely hampered by the absence of any specific high-affinity inhibitor. Here we describe advanced procedures for purifying a candidate inhibitor, previously called endogenous inhibitor factor (NCX(IF)), and demonstrate its direct actions on NCX1 activities in the single-cell system. A combination of advanced HILIC (hydrophilic interaction liquid chromatography) procedures with analytical tests suggests that the properties of NCX(IF) resemble those of a small (disaccharide size) polar molecule lacking any aromatic rings, conjugated bonds, or a primary amino group. The effects of NCX(IF) on the NCX1-mediated ion currents (I(NCX)) and cytosolic Ca(2+) extrusion were detected by a combination of patch-clamp and confocal microscopy under conditions in which the purified NCX(IF) was directly loaded into the cytoplasm of patched cardiomyocytes. It was demonstrated that cytosolic NCX(IF) blocks the Ca(2+)-activated NCX1 inward current and the accompanying extrusion of Ca(2+) from the cell with high efficacy. A constant fraction of NCX1 inhibition was observed under conditions in which the cytosolic [Ca(2+)](i) was varied at fixed doses of NCX(IF), suggesting that the degree of inhibition is controlled by NCX(IF) dose and not by cytosolic Ca(2+) concentration. NCX(IF) blocks equally well both the Ca(2+) extrusion and Ca(2+) entry modes of NCX1, consistent with thermodynamic principles expected for the functioning of a bidirectional carrier-type transport system. We concluded that NCX(IF) interacts with a putative regulatory domain from the cytosolic side and, thus, may play an important regulatory role in controlling Ca(2+) signaling in the heart. This may represent a new potential tool for developing novel treatments for cardiac Ca(2+) signaling dysfunction." @default.
• W2162078918 created "2016-06-24" @default.
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• W2162078918 date "2008-05-29" @default.
• W2162078918 modified "2023-09-27" @default.
• W2162078918 title "Direct Loading of the Purified Endogenous Inhibitor into the Cytoplasm of Patched Cardiomyocytes Blocks the Ion Currents and Calcium Transport through the NCX1 Protein" @default.
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• W2162078918 doi "https://doi.org/10.1021/bi8004279" @default.
• W2162078918 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/3522518" @default.
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