Guanine Nucleotide Exchange Assay Using Fluorescent MANT-GDP
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Updated: Feb 8, 2026

Use of Stopped-Flow Fluorescence and Labeled Nucleotides to Analyze the ATP Turnover Cycle of Kinesins
Published on: October 17, 2014
Tomoharu Kanie1, Peter K Jackson1
1Baxter Laboratory, Department of Microbiology & Immunology and Department of Pathology, Stanford University School of Medicine, Stanford, CA, USA.
Abstract:
GTPases are molecular switches that cycle between the inactive GDP-bound state and the active GTP-bound state. GTPases exchange nucleotides either by its intrinsic nucleotide exchange or by interaction with guanine nucleotide exchange factors (GEFs). Monitoring the nucleotide exchange in vitro, together with reconstitution of direct interactions with regulatory proteins, provides key insights into how a GTPase is activated. In this protocol, we describe core methods to monitor nucleotide exchange using fluorescent N-Methylanthraniloyl (MANT)-guanine nucleotide.
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