Localization of (Ca2+ + Mg2+)-ATPase,Ca2+ pump and other ATPase activities in cardiac sarcolemma |
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Authors: | NCharle Morcos |
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Institution: | Department of Physiology, and the American Heart Association, Greater Los Angeles Affiliate, Cardiovascular Research Laboratories, UCLA School of Medicine, Los Angeles, CA 90024 U.S.A. |
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Abstract: | N-Ethylmaleimide was employed as a surface label for sarcolemmal proteins after demonstrating that it does not penetrate to the intracellular space at concentrations below 1·10?4 M. The sarcolemmal markers, ouabain-sensitive (Na+ + K+)-ATPase and Na+/Ca2+-exchange activities, were inhibited in N-ethylmaleimide perfused hearts. Intracellular activities such as creatine phosphokinase, glutamate-oxaloacetate transaminase and the internal phosphatase site of the Na+ pump (K+-p-nitrophosphatase) were not affected. Almost 20% of the (Ca2+ + Mg2+)-ATPase and Ca2+ pump were inhibited indicating the localization of a portion of this activity in the sarcolemma. Sarcolemma purified by a recent method (Morcos, N.C. and Drummond, G.I. (1980) Biochim. Biophys. Acta 598, 27–39) from N-ethylmaleimide-perfused hearts showed loss of approx. 85% of its (Ca2+ + Mg2+-ATPase and Ca2+ pump compared to control hearts. (Ca2+ + Mg2+)-ATPase and Ca2+ pump activities showed two classes of sensitivity to vanadate ion inhibition. The high vanadate affinity class ( for inhibition approx. 1.5 μM) may be localized in the sarcolemma and represented approx. 20% of the total inhibitable activity in agreement with estimates from N-ethylmaleimide studies. Sucrose density fractionation indicated that only a small portion of Mg2+-ATPase and Ca2+-ATPase may be associated with the sarcolemma. The major portion of these activities seems to be associated with high density particles. |
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Keywords: | Enzyme localization (Cardiac sarcolemma) EGTA Address for correspondence: Cardiovascular Research Laboratory A3-381 CHS UCLA School of Medicine Los Angeles CA 90024 U S A |
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