Quantification of GTPase Cycling Rates of GTPases and GTPase:Effector Mixtures Using GTPase Glo Assays

Sophie Tschirpke, Werner K.G. Daalman, Liedewij Laan*

*Corresponding author for this work

Research output: Contribution to journalArticleScientificpeer-review

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.

Original languageEnglish
Article numbere1000
Number of pages29
JournalCurrent Protocols
Volume4
Issue number4
DOIs
Publication statusPublished - 2024

Keywords

  • Cdc42
  • GTPase activity
  • GTPase assay
  • GTPase cycling rates
  • GTPase effectors

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