VpAAT1, a Gene Encoding an Alcohol Acyltransferase, Is Involved in Ester Biosynthesis during Ripening of Mountain Papaya Fruit

Abstract

Mountain papaya ( Vasconcellea pubescens ) is a climacteric fruit that develops a strong and characteristic aroma during ripening. Esters are the main volatile compounds produced by the fruit, and most of them are dependent on ethylene. As esters are synthesized through alcohol acyltransferases (AAT), a full-length cDNA (VpAAT1) was isolated that displayed the characteristic motifs of most plant acyltransferases. The full-length cDNA sequence was cloned and expressed in yeasts, obtaining a functional enzyme with high AAT activity toward the formation of benzyl acetate. The transcript accumulation pattern provided by qPCR analysis showed that the VpAAT1 gene is expressed exclusively in fruit tissues and that a high level of transcripts is accumulated during ripening. The increase in VpAAT1 transcripts in fruit is coincident with the increase in AAT activity; transcript accumulation is induced by ethylene, and it is avoided by 1-methylcyclopropene (1-MCP) treatment. The data indicate that VpAAT1 is involved in aroma formation and that ethylene plays a major role in regulating its expression.

Cloning and functional characterization of two monoterpene acetyltransferases from glandular trichomes of L. x intermedia.

“…Kinetic parameters of LiAAT-3 and LiAAT-4 are in the range of those reported for previously cloned or biochemically characterized alcohol acyltransferase genes from a number of different plants (Aharoni et al 2000;Beekwilder et al 2004;ElSharkawy et al 2005;Shalit et al 2003). In addition, enzymatic activity of FvVAAT, CmAAT-4, CbBEBT, VpAAT1 for geraniol and nerol were reported to be in the range of 10-3451 pkat/mg in the presence of acetyl CoA (Balbontin et al 2010;Beekwilder et al 2004;D'Auria et al 2002;El-Sharkawy et al 2005).…”

Computational study enlightens the structural role of the alcohol acyltransferase DFGWG motif

“…Interestingly, a decrease in enzyme activity was reported for Y to F substitution of this conserved residue [28]. This residue is not conserved in Musa sapienta AAT, a close relative to VpAAT1, which displays a Phe residue instead [4]. A recent publication showed a similar result giving insights into the role of Y40 (Y52 in VpAAT1) based on The box indicates the AFGWG motif and the black arrow the loop that moves towards the center of the solvent channel.…”

“…VpAAT1 enzyme has a strong activity to produce benzyl acetate [4], showing the best affinity energy for benzyl alcohol and acetyl-CoA [15]. The interaction of these substrates with the active site of VpAAT1 mutants was studied using MDS (Fig.…”

“…Vasconcellea pubescens alcohol acyltransferase (VpAAT1)-a member of the BAHD superfamily-was isolated and characterized from ripe mountain papaya fruit [4,5]. VpAAT1 belongs to subfamily III [4], where other AATs that take part in the synthesis of volatile compounds during fruits ripening such as melon [6] and apple [7][8][9] are grouped. A low sequence similarity can be observed for these proteins; nevertheless, similar acyltransferase function and the presence of two highly conserved motifs are found [10].…”

“…However, phylogenetically closely related enzymes, especially those responsible for volatile ester formation, can share up to 90 % identity. In fact, 55-68 % sequence identity is reported for AATs clustered in subgroup III [4]. Recently, in VpAAT1, the substitution of histidine and aspartic acid residues from the HxxxD motif showed that both residues are crucial for activity, H166 being directly involved in catalysis while D170 plays a structural role in the maintenance of this channel structure [11].…”