Determination of coupling ratios of the calcium pump of sarcoplasmic reticulum by pulse methods.
Coupling of Ca2+ transport to ATP hydrolysis in isolated sarcoplasmic reticulum vesicles has been studied following pulsed additions of either ATP or Ca2+. ATP was infused as a pulse into medium, whose free Ca2+ concentration was maintained constant at saturating levels by a calciumstat procedure, using either a Ca2+-selective electrode or the spectrophotometric arsenazo III technique as Ca2+ indicators. The low ATP levels virtually exclude contributions by "basal" ATPase activity. Passive leakage of Ca2+, monitored after an ATP pulse, does not contribute more than 5% to subintegral coupling ratios. Pulsed additions of Ca2+ were made into medium. containing saturating concentrations of ATP, whose hydrolysis was monitored by a pH-stat procedure. Ca2+-stimulated hydrolysis continued until all the Ca2+ was transported into the vesicles. Values for the coupling ratio, Ca2+/ATP, of 1.82 +/- 0.12 and 1.79 +/- 0.15 were obtained by the ATP- and Ca2+-pulse methods, respectively.
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001150:Arsenazo III; D001693:Biological Transport, Active / Q000187:drug effects; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D004566:Electrodes; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D066298:In Vitro Techniques; D008722:Methods; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
D001150:Arsenazo III; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases; D002118:Calcium