Abstract
Terpenes are one of the largest and most diversified families of natural compounds. Although they have found numerous industrial applications, the molecular basis of their synthesis in plants has, until now, not been fully understood. Plant genomes have been shown to contain dozens of terpene synthase (TPS) genes, however knowledge of their amino-acidic protein sequence in not sufficient to predict which terpene(s) will be produced by a particular enzyme. In order to investigate the structural basis of a TPS specificity, we performed site directed mutations in the geraniol synthase from Ocimum basilicum. The results obtained suggest that a specific region on the catalytic site plays an important role in GPP transformation, either by stabilizing the GPP substrate on the catalytic site, or by enabling its transformation into a monoterpenol via an intermediate carbocation.
Highlights
The immense variety of chemical compositions encountered in aromatic plants has always attracted human interest, and such plants are believed to have been cultivated since the Neolithic for their fragrance, food improvement abilities, and traditional medicinal properties
The product specificities of monoterpenol synthases have been extensively studied using heterologous expression, the manner in which their amino acidic sequence relates to the specific enzymatic function still remains largely unknown, with the notable exception of the sabinene and 1,8 cineole synthases from S. fruticosa [11]
To enable catalytic site analysis, we positioned each residue differing from O. basilicum and P. frutescens geraniol synthase (GES) and LIS within the predicted catalytic site, and used an in vivo characterization approach to investigate the functional consequences of amino-acidic substitutions at the putative catalytic site of O. basilicum GES
Summary
The immense variety of chemical compositions encountered in aromatic plants has always attracted human interest, and such plants are believed to have been cultivated since the Neolithic for their fragrance, food improvement abilities, and traditional medicinal properties. Monoterpene aroma are amongst the most basic and prevalent fragrance representatives They belong to the family of plant isoprenoids, which are amongst the most diversified compounds, with over 40,000 molecules having been described [1]. The tremendous range of possible variations in the carbocationic reactions (cyclizations, hydride shifts, rearrangements, and termination steps) catalyzed by terpenoid synthases explains the wide range of possible products [6]. Such studies have usually been carried out using heterologous truncated cDNA expression in Escherichia coli (since the native plant enzyme targets the chloroplast and bacteria cannot process introns), followed by enzymatic assays with crude cell extracts
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