Monomeric solubilized sarcoplasmic reticulum Ca pump protein: demonstration of Ca binding and dissociation coupled to ATP hydrolysis.
The sarcoplasmic reticulum Ca-pump protein can be solubilized in monomeric form in nonionic detergents with full retention of ATPase activity. It is impossible to prove directly that coupling between ATP hydrolysis and Ca2+ transport is maintained in the soluble state, because separate compartments for Ca2+ uptake and Ca2+ discharge are required to demonstrate this, but here we provide strong indirect evidence that coupling in fact persists, in both the forward and reverse directions of the normal pump cycle. Demonstration of coupling in the forward direction makes use of the fact that the solubilized protein is structurally labile in the absence of bound Ca2+. Loss of activity accompanying ATP hydrolysis in solution is quantitatively consistent with sequential Ca2+ binding and dissociation during the hydrolysis cycle. Coupling in the reverse direction is demonstrated by the dependence of ATP synthesis on Ca2+ concentration. Although substantial differences between solubilized (monomeric) and membrane-bound protein are shown to exist, as previously reported, they do not affect the main conclusion from this work, which is that the molecular machinery for free-energy coupling in active Ca2+ transport is an inherent property of each individual catalytic polypeptide chain of the pump protein.
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003546:Cysteine Endopeptidases; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D012519:Sarcoplasmic Reticulum / Q000032:analysis*; D012995:Solubility; D013816:Thermodynamics
D007473:Ion Channels; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; C043440:arginyllysine esteropeptidase; D000252:Calcium-Transporting ATPases; D002118:Calcium