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GTPases and their Regulation

GTPases and their Regulation

Guanine nucleotide-binding proteins (G-proteins), also known as GTPases, are a superfamily of proteins that regulate many cellular processes, such as cell signaling, vesicular transport, and the regulation of cell shape and motility. Mutation or dysfunction of these proteins can lead to disease. There are around 40,000 known G-proteins that can broadly be classified into two groups ‒ small G-proteins consisting of a single domain and large multi-domain G-proteins.
Large G-proteins,...

Activation and Inactivation of G Proteins

Activation and Inactivation of G Proteins

Heterotrimeric G proteins are guanine nucleotide-binding proteins. As the name suggests, heterotrimeric G proteins are composed of three subunits: alpha, beta, and gamma. They remain GDP-bound or GTP-bound inside the cells and switch between inactive/active states. The Gα subunit possesses the nucleotide-binding pocket that binds guanine nucleotides and switches between GDP or GTP-bound states. In contrast, the Gꞵ and Gγ subunits are always bound together with high...

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Related Experiment Video

Updated: Apr 17, 2026

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

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

Published on: November 11, 2018

GTP-specific fab fragment-based GTPase activity assay.

Kari Kopra¹, Anita Rozwandowicz-Jansen, Markku Syrjänpää

¹†Institute of Biomedicine, Department of Cell Biology and Anatomy, University of Turku, Kiinamyllynkatu 10, Third Floor, FI-20520 Turku, Finland.

Analytical Chemistry

|February 25, 2015

Summary

Researchers developed a novel GTP-specific antibody fragment to detect free guanosine triphosphate (GTP). This tool enables sensitive monitoring of GTP levels and GTPase activity in cellular signaling pathways.

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Last Updated: Apr 17, 2026

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

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Published on: November 11, 2018

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Published on: August 22, 2010

Area of Science:

Biochemistry
Molecular Biology
Cellular Signaling

Background:

GTPases are crucial for cellular signaling, cycling between inactive (GDP-bound) and active (GTP-bound) states.
Ras GTPases are frequently altered in cancer, yet effective inhibitors are limited.
Current methods for monitoring GTP hydrolysis often use luminescent analogs.

Purpose of the Study:

To develop and characterize a novel GTP-specific antibody fragment.
To create a sensitive assay for monitoring GTP concentration and GTPase activity.

Main Methods:

Phage display technology was used to select antibody fragments against GTP-biotin conjugates.
The most potent fragment, 2A4(GTP), was identified and characterized for its specificity.
A heterogeneous time-resolved luminescence assay was developed using the 2A4(GTP) fragment.

Main Results:

Six Fab fragments with specificity for GTP were selected.
The 2A4(GTP) fragment demonstrated over 100-fold specificity for GTP compared to GDP, ATP, or CTP.
The developed assay can monitor GTP concentration and GTPase activity (H-Ras activation/deactivation) at nanomolar protein levels.

Conclusions:

A novel, highly specific GTP-binding Fab fragment (2A4(GTP)) was successfully developed.
This fragment enables a sensitive luminescence-based assay for studying GTPase function.
The assay facilitates research into cellular signaling pathways involving GTPases, particularly Ras proteins.