Protocol for structural and biochemical analyses of RhoA GTPase

STAR Protoc. 2021 May 14;2(2):100541. doi: 10.1016/j.xpro.2021.100541. eCollection 2021 Jun 18.

Abstract

Ras GTPases in complex with Guanosine triphosphate (GTP) or GTP analog exhibit dynamic equilibrium between two interconvertible conformations-an inactive state 1 and an active state 2. Unlike Ras, it remains unclear if the GTP-bound form of Rho GTPases also exhibits multiple conformational states. Here, we describe a protocol for structural and biochemical analyses of RhoA GTPase. This protocol can be adapted for the characterization of other Rho GTPases. For details on the use and execution of this protocol, please refer to Lin et al. (2021).

Keywords: NMR; Protein expression and purification; Signal Transduction; Structural Biology; X-ray Crystallography.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Escherichia coli
  • Guanosine Triphosphate / chemistry
  • Guanosine Triphosphate / metabolism
  • Humans
  • Models, Molecular
  • Protein Conformation
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • rho GTP-Binding Proteins* / chemistry
  • rho GTP-Binding Proteins* / genetics
  • rho GTP-Binding Proteins* / isolation & purification
  • rho GTP-Binding Proteins* / metabolism
  • rhoA GTP-Binding Protein / chemistry
  • rhoA GTP-Binding Protein / genetics
  • rhoA GTP-Binding Protein / isolation & purification
  • rhoA GTP-Binding Protein / metabolism

Substances

  • Recombinant Proteins
  • RHOA protein, human
  • Guanosine Triphosphate
  • rho GTP-Binding Proteins
  • rhoA GTP-Binding Protein