Abstract
ATP provides the energy that is essential for all P-type ATPases to actively transport their substrates against an existing gradient. This ATP hydrolysis can be measured using different methods. Here, we describe a method that uses radiolabeled [γ-32P]ATP, which is hydrolyzed by P-type ATPases to ADP and 32Pi. Activated charcoal is used to bind the excess of [γ-32P]ATP, which can be separated from the unbound 32Pi by centrifugation. With this method, a wide range (0.1 μM–10 mM) of ATP can be used. In addition, we also describe in detail how ATP hydrolysis is translated into ATPase activity.
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Swarts, H.G.P., Koenderink, J.B. (2016). ATPase Activity Measurements Using Radiolabeled ATP. In: Bublitz, M. (eds) P-Type ATPases. Methods in Molecular Biology, vol 1377. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3179-8_13
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DOI: https://doi.org/10.1007/978-1-4939-3179-8_13
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3178-1
Online ISBN: 978-1-4939-3179-8
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