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Licensed Unlicensed Requires Authentication Published by De Gruyter November 24, 2005

Monitoring the real-time kinetics of the hydrolysis reaction of guanine nucleotide-binding proteins

  • Alexander Eberth , Radovan Dvorsky , Christian F.W. Becker , Andrea Beste , Roger S. Goody and Mohammad Reza Ahmadian
From the journal Biological Chemistry

Abstract

The conversion of guanosine triphosphate (GTP) to guanosine diphosphate (GDP) and inorganic phosphate (Pi) by guanine nucleotide-binding proteins (GNBPs) is a fundamental enzyme reaction in living cells that acts as an important timer in a variety of biological processes. This reaction is intrinsically slow but can be stimulated by GTPase-activating proteins (GAPs) by several orders of magnitude. In the present study, we synthesized and characterized a new fluorescent nucleotide, 2′(3′)-O-(N-ethylcarbamoyl-(5″-carboxytetramethylrhodamine) amide)-GTP, or tamraGTP, which is sensitive towards conformational changes of certain GNBPs induced by GTP hydrolysis. Unlike other fluorescent nucleotides, tamra-GTP allows real-time monitoring of the kinetics of the intrinsic and GAP-catalyzed GTP hydrolysis reactions of small GNBPs from the Rho family.

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Corresponding author

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Published Online: 2005-11-24
Published in Print: 2005-11-01

©2005 by Walter de Gruyter Berlin New York

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