Characterization of a high-affinity Mg2+-independent Ca2+-ATPase from rat brain synaptosomal membranes.
A high-affinity Mg2+-independent Ca2+-ATPase (Ca2+-ATPase) has been differentiated from the Mg2+-dependent, Ca2+-stimulated ATPase (Ca2+,Mg2+-ATPase) in rat brain synaptosomal membranes. Using ATP as a substrate, the K0.5 of Ca2+ for Ca2+-ATPase was found to be 1.33 microM with a Km for ATP of 19 microM and a Vmax of 33 nmol/mg/min. Using Ca-ATP as a substrate, the Km for Ca-ATP was found to be 0.22 microM. Unlike Ca2+,Mg2+-ATPase, Ca2+-ATPase was not inhibited by N-ethylmaleimide, trifluoperazine, lanthanum, zinc, or vanadate. La3+ and Zn2+, in contrast, stimulated the enzyme activity. Unlike Ca2+, Mg2+-ATPase activity, ATP-dependent Ca2+ uptake was negligible in the absence of added Mg2+, indicating that the Ca2+ transport into synaptosomal endoplasmic reticulum may not be a function of the Ca2+-ATPase described. Ca2+-ATPase activity was not stimulated by the monovalent cations Na+ or K+. Ca2+, Mg2+-ATPase demonstrated a substrate preference for ATP and ADP, but not GTP, whereas Ca2+-ATPase hydrolyzed ATP and GTP, and to a lesser extent ADP. The results presented here suggest the high-affinity Mg2+-independent Ca2+-ATPase may be a separate form from Ca2+,Mg2+-ATPase. The capacity of Mg2+-independent Ca2+-ATPase to hydrolyze GTP suggests this protein may be involved in GTP-dependent activities within the cell.
D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D001693:Biological Transport, Active / Q000187:drug effects; D001921:Brain / Q000201:enzymology*; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins / Q000494:pharmacology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D008297:Male; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D012964:Sodium / Q000494:pharmacology; D013379:Substrate Specificity; D013574:Synaptosomes / Q000201:enzymology*; D013861:Thiocyanates / Q000494:pharmacology
D005452:Fluoresceins; D013861:Thiocyanates; D010042:Ouabain; D000255:Adenosine Triphosphate; D012964:Sodium; D017301:Ca(2+) Mg(2+)-ATPase; D000252:Calcium-Transporting ATPases; D016650:Fluorescein-5-isothiocyanate; D008274:Magnesium; D011188:Potassium; D002118:Calcium