Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay

Javad Alizadeh , Shahla Shojaei , Simone da Silva Rosa , Adel Rezaei Moghadam , Amir Zeki , Mohammad Hashemi , Marek Los (1, 2) , Joseph W Gordon , Saeid Ghavami

1 CBM - Centre de biophysique moléculaire
2 Le Studium Loire Valley Institute for Advanced Studies, 45000 Orléans, France

The Rho GTPase family belongs to the Ras superfamily and includes approximately 20 members in humans. Rho GTPases are important in the regulation of diverse cellular functions, including cytoskeletal dynamics, cell motility, cell polarity, axonal guidance, vesicular trafficking, and cell cycle control. Changes in Rho GTPase signaling play an essential regulatory role in many pathological conditions, such as cancer, central nervous system diseases, and immune system-dependent diseases. The posttranslational modification of Rho GTPases (i.e., prenylation by mevalonate pathway intermediates) and GTP binding are key factors which affect the activation of this protein. In this paper, two essential and simple methods are provided to detect a broad range of Rho GTPase prenylation and GTP binding activities. Details of the technical procedures that have been used are explained step by step in this manuscript.

Mots clés: Biochemistry; Issue 141; Simvastatin; glioblastoma; Rho GTPase; cancer cell biology; prenylation; mevalonate pathway

Domaines: Sciences du Vivant [q-bio]; Cancer; Biochimie, Biologie Moléculaire

Javad Alizadeh, Shahla Shojaei, Simone da Silva Rosa, Adel Rezaei Moghadam, Amir Zeki, et al.. Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay. Journal of visualized experiments : JoVE, 2018, 141, ⟨10.3791/57646⟩. ⟨hal-02506767⟩