Formation of a stable inactive complex of the sarcoplasmic reticulum calcium ATPase with magnesium, beryllium, and fluoride.
Incubation of leaky or nonionic detergent-solubilized sarcoplasmic reticulum vesicles in solutions containing magnesium, beryllium, and fluoride caused a time-dependent complete inhibition of calcium ATPase activity. The inhibited state persisted through dialysis for 2 days versus EGTA and was reversed within minutes by the presence of 0.5 mM calcium. Calcium-independent ATPase activity was unaffected. Omission of magnesium or fluoride resulted in retention of activity, while omission of beryllium produced slower inactivation, as described previously (Murphy, A. J., and Coll, R. J. (1992) J. Biol. Chem. 267, 5229-5235). Incubation of nonleaky vesicles had a similar effect, although it occurred more than 10-fold more slowly, suggesting that a component, probably beryllium, must enter the vesicles for inhibition to occur. By contrast, inhibition of nonleaky vesicles by magnesium and fluoride developed less than 2-fold more slowly. Including calcium in the incubation mixtures resulted in partial protection, so that the time course of CaATPase activity leveled off at nonzero values (for example, at 0.1 mM calcium, the activity leveled off at 43% of control). This result is most simply accounted for by a model involving simultaneous binding of calcium and a form of fluoroberyllium to the CaATPase (e.g. Ca2EMgBeFn).
Murphy
A J
AJ
Department of Biochemistry, School of Dentistry, University of the Pacific, San Francisco, California 94115.
Coll
R J
RJ
D000244:Adenosine Diphosphate / Q000378:metabolism; D000818:Animals; D001608:Beryllium / Q000737:chemistry*; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors* / Q000737:chemistry; D005459:Fluorides / Q000737:chemistry*; D066298:In Vitro Techniques; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry*; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*
D000001:Calcimycin; D000244:Adenosine Diphosphate; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D001608:Beryllium; D005459:Fluorides; D002118:Calcium