title: Fluoride binding to the calcium ATPase of sarcoplasmic reticulum converts its transport sites to a low affinity, lumen-facing form.
abstract: The sarcoplasmic reticulum CaATPase forms an inactive complex with fluoride (CaATPase-F), which in the absence of calcium reactivates very slowly (t1/2 approximately 40 h at 25 degrees C). Reactivation is greatly accelerated (greater than 10(3)) by calcium in the millimolar range provided it has access to luminal sites of the enzyme. Measurement of the calcium concentration dependence of the reactivation rate constant revealed a saturable effect with a midpoint of about 12 mM calcium. These results show that an effect other than phosphorylation can produce a greater than 10(3)-fold affinity decrease and reorientation of the calcium transport sites. At a fixed calcium concentration, reactivation became faster with increasing pH (pKa greater than 8), suggesting competition between Ca2+ and H+ for transport sites. CaATPase-F lacked the ability to bind calcium with high affinity or to form phosphorylated enzyme intermediate from Pi; it bound adenyl-5'-yl methylenediphosphonate more than 10-fold less strongly than control CaATPase, had numerous sulfhydryl groups with significantly different reactivity, and was notably less susceptible (more than 10-fold) to thermal inactivation compared with control Ca-ATPase. These results suggest that formation of Ca-ATPase-F involves significant structural changes.
authors.affiliation: Department of Biochemistry, School of Dentistry, University of the Pacific, San Francisco, California 94115.
authors.forename: A J
authors.initials: AJ
authors.lastname: Murphy
authors.forename: R J
authors.initials: RJ
authors.lastname: Coll
mesh_terms: D000818:Animals; D001665:Binding Sites; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005459:Fluorides / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D011487:Protein Conformation; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013816:Thermodynamics; D013997:Time Factors
chemical_list: D000252:Calcium-Transporting ATPases; D005459:Fluorides