Quantification of GTPase Cycling Rates of GTPases and GTPase:Effector Mixtures Using GTPase Glo Assays
Sophie Tschirpke (TU Delft - BN/Liedewij Laan Lab)
Werner K.G. Daalman (Student TU Delft)
L. Laan (TU Delft - BN/Liedewij Laan Lab)
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Abstract
In different cellular activities such as signal transduction, cell division, and intracellular transportation, small guanosine triphosphatases (GTPases) take on a vital role. Their function involves hydrolysis of guanosine triphosphate (GTP) to guanosine diphosphate (GDP). In this article, we explain the application of a commercially available GTPase assay—the GTPase Glo assay by Promega—for investigation of GTPase-effector interactions. We provide experimental protocols together with an analysis model and software to obtain GTPase cycling rates of GTPases and GTPase:effector mixtures. GTPase cycling rates refer to the rates by which a GTPase completes an entire GTPase cycle. These rates enable quantification of the strength of GTPase effectors in a concentration-dependent fashion, as well as quantification of the combined effect of two effectors, independent of which GTPase cycle step they are affecting.
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