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Molecular & Cellular Proteomics 4:936-944, 2005.
© 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

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Research

Biochemical Clustering of Monomeric GTPases of the Ras Superfamily ^*,S

Marie-Elaine Caruso^,, Sarah Jenna^,¶,||, Simon Beaulne, Eun-Hye Lee^¶,||, Anne Bergeron^**, Cédric Chauve^**, Philippe Roby, Jean-François Rual, David E. Hill, Marc Vidal, Roger Bossé^,¶¶ and Eric Chevet^,,¶,||||

From the Organelle Signaling Laboratory, Department of Surgery and ^¶ Montreal Proteomics Network, McGill University, Montreal, Quebec H3A 2B2, Canada, the ^** Laboratoire de Combinatoire et d’Informatique Mathématique, University of Quebec at Montreal, Montreal, Quebec H3C 3P8, Canada, PerkinElmer BioSignal, Montreal, Quebec H3J 1R4, Canada, and the Centre for Cancer Systems Biology and Department of Cancer Biology, Dana-Farber Cancer Institute and the Department of Genetics, Harvard Medical School, Boston, Massachusetts 02115

To date phylogeny has been used to compare entire families of proteins based on their nucleotide or amino acid sequence. Here we developed a novel analytical platform allowing a systematic comparison of protein families based on their biochemical properties. This approach was validated on the Rho subfamily of GTPases. We used two high throughput methods, referred to as AlphaScreen^TM and FlashPlate®, to measure nucleotide binding capacity, exchange, and hydrolysis activities of small monomeric GTPases. These two technologies have the characteristics to be very sensitive and to allow homogenous and high throughput assays. To analyze and integrate the data obtained, we developed an algorithm that allows the classification of GTPases according to their enzymatic activities. Integration and hierarchical clustering of these results revealed unexpected features of the small Rho GTPases when compared with primary sequence-based trees. Hence we propose a novel phylobiochemical classification of the Ras superfamily of GTPases.

^|||| To whom correspondence may be addressed. Tel.: 514-934-1934 (ext. 35468); Fax: 514-843-1411; E-mail: eric.chevet{at}mcgill.ca

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