Utilization of X-537A to distinguish between intravesicular and membrane-bound calcium ions in sarcoplasmic reticulum.
The Ca2+ ionophore X-537A is employed as a tool to distinguish between intravesicular Ca2+ and surface membrane-bound Ca2+ in sarcoplasmic reticulum isolated from rabbit skeletal muscle. When sarcoplasmic reticulum is incubated in 20 mM Ca2+ in the absence of ATP, 10-12 h are necessary for measurable amount of Ca2+ to penetrate into the vesicular space, as determined by the fact that X-537A releases Ca2+ from 'loaded' vesicles only after this period of incubation. A fraction of Ca2+ of 50-60 nmol/mg protein, rapidly taken up by sarcoplasmic reticulum, exchanges with Mg2+ and K+ in the medium and is readily released by ethyleneglycol-bis-(beta-aminoethyl ether)-N,N'-tetraacetic acid, but it is not released by X-537A. The slow-penetrating fraction of Ca2+ (30-40 nmol/mg protein) is rapidly released X-537A. The results indicate that most of the Ca2+ retained by sarcoplasmic reticulum under conditions of passive uptake is bound to the external side of the membrane. The fraction of Ca2+ that slowly penetrates the vesicles remains essentially free inside the vesicles and only a small part is bound to the internal side of the membrane.
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D007832:Lasalocid / Q000494:pharmacology*; D008274:Magnesium / Q000378:metabolism; D008566:Membranes / Q000378:metabolism*; D011817:Rabbits; D011955:Receptors, Drug / Q000187:drug effects*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D011955:Receptors, Drug; D008274:Magnesium; D002118:Calcium; D007832:Lasalocid