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Functional characterization of a melon alcohol acyl-transferase gene family involved in the biosynthesis of ester volatiles. Identification of the crucial role of a threonine residue for enzyme activity

El-Sharkawy, Islam and Manriquez, Daniel and Flores, Francisco B. and Regad, Farid and Bouzayen, Mondher and Latché, Alain and Pech, Jean-Claude Functional characterization of a melon alcohol acyl-transferase gene family involved in the biosynthesis of ester volatiles. Identification of the crucial role of a threonine residue for enzyme activity. (2005) Plant Molecular Biology, vol. 5 (n° 2). pp. 345-362. ISSN 0167-4412

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Official URL: http://dx.doi.org/10.1007/s11103-005-8884-y

Abstract

Volatile esters, a major class of compounds contributing to the aroma of many fruit, are synthesized by alcohol acyl-transferases (AAT). We demonstrate here that, in Charentais melon (Cucumis melo var. cantalupensis), AAT are encoded by a gene family of at least four members with amino acid identity ranging from 84% (Cm-AAT1/Cm-AAT2) and 58% (Cm-AAT1/Cm-AAT3) to only 22% (Cm-AAT1/Cm-AAT4). All encoded proteins, except Cm-AAT2, were enzymatically active upon expression in yeast and show differential substrate preferences. Cm-AAT1 protein produces a wide range of short and long-chain acyl esters but has strong preference for the formation of E-2-hexenyl acetate and hexyl hexanoate. Cm-AAT3 also accepts a wide range of substrates but with very strong preference for producing benzyl acetate. Cm-AAT4 is almost exclusively devoted to the formation of acetates, with strong preference for cinnamoyl acetate. Site directed mutagenesis demonstrated that the failure of Cm-AAT2 to produce volatile esters is related to the presence of a 268-alanine residue instead of threonine as in all active AAT proteins. Mutating 268-A into 268-T of Cm-AAT2 restored enzyme activity, while mutating 268-T into 268-A abolished activity of Cm-AAT1. Activities of all three proteins measured with the prefered substrates sharply increase during fruit ripening. The expression of all Cm-AAT genes is up-regulated during ripening and inhibited in antisense ACC oxidase melons and in fruit treated with the ethylene antagonist 1-methylcyclopropene (1-MCP), indicating a positive regulation by ethylene. The data presented in this work suggest that the multiplicity of AAT genes accounts for the great diversity of esters formed in melon.

Item Type:Article
Additional Information:Thanks to Springer editor.The definitive version is available at www.springerlink.com . The original PDF of the article can be found at Plant Molecular Biology website : http://www.springerlink.com/content/0167-4412
Audience (journal):International peer-reviewed journal
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Institution:French research institutions > Institut National de la Recherche Agronomique - INRA
Université de Toulouse > Institut National Polytechnique de Toulouse - INPT
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Deposited By: Dabernat Séverine

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