Abstract
Large amounts of enzyme activity are often lost, when membrane preparations are treated with detergents for solubilisation of E-type ATPases. In this work, however, it is shown, that 1) detergents do not inactivate the ecto-ATPase activity of intact rat tissues (mesenteric artery, ductus deferens and lung). 2) Neither detergents nor ATP gave rise to release of ATPase activity when administered separately to the tissue incubation, but when simultaneously present, detergent and ATP caused the ATPase activity to be released to the medium. The released ATPase activity 3) stayed in the supernatant upon ultracentrifugation and 4) showed the characteristics of E-type ATPases. 5) Several nucleotides, including nonhydrolysable analogues, could replace ATP mediating extraction of active E-type ATPase with detergents. 6) With an average yield of 3 U per gram of tissue, nucleotide/detergent extraction may provide a useful method for obtaining a solubilized form of E-type ATPase (specific activity: 0.3, 0.1 and 0.03 U per mg of protein in extracts from mesenteric artery, ductus deferens and lung, respectively).
The experiments are interpreted to indicate that in intact cells, nucleotides induce a modification of the protein(s) that hydrolyze extracellular ATP. The modified protein is released from the cell membrane in a hydrolytically active form in the presence of detergents. The modification requires contact with the rest of the cell, since nucleotides do not protect the E-type activity of membrane preparations during detergent treatment, i.e. the modification does not occur just because the enzyme is binding or hydrolyzing its substrate.
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© 1997 Springer Science+Business Media New York
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Plesner, L. (1997). Solubilized E-Type ATPase is Released from Intact Rat Tissues in the Simultaneous Presence of Nucleotides and Detergents. In: Plesner, L., Kirley, T.L., Knowles, A.F. (eds) Ecto-ATPases. Springer, Boston, MA. https://doi.org/10.1007/978-1-4615-5955-9_6
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DOI: https://doi.org/10.1007/978-1-4615-5955-9_6
Publisher Name: Springer, Boston, MA
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