Predicting the substrate specificity of a glycosyltransferase implicated in the production of phenolic volatiles in tomato fruit

• Published in: The FEBS journal 2011-01, Vol.278 (2), p.390-400
• Main Authors: Louveau, Thomas, Leitao, Celine, Green, Sol, Hamiaux, Cyril, van der Rest, Benoît, Dechy-Cabaret, Odile, Atkinson, Ross G, Chervin, Christian
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; aroma; Benzyl Alcohol - chemistry; Benzyl Alcohol - metabolism; Benzyl Alcohols - chemistry; Benzyl Alcohols - metabolism; Biochemistry; Catalytic Domain - physiology; Chromosomes - genetics; Cloning, Molecular; docking; Environmental Sciences; Escherichia coli - genetics; Escherichia coli - metabolism; eugenol; Eugenol - chemistry; Eugenol - metabolism; Gene expression; Gene Expression - genetics; Glycosylation; Glycosyltransferases - chemistry; Glycosyltransferases - genetics; Glycosyltransferases - metabolism; guaiacol; Guaiacol - chemistry; Guaiacol - metabolism; Humanities and Social Sciences; Hydrogen Bonding; Hydrogen bonds; Hydroquinones - chemistry; Hydroquinones - metabolism; isosalicin; Life Sciences; Lycopersicon esculentum - enzymology; Lycopersicon esculentum - genetics; methyl salicylate; Models, Molecular; Molecular Sequence Data; Phenols - metabolism; Phenylethyl Alcohol - chemistry; Phenylethyl Alcohol - metabolism; Phylogeny; Plant Proteins - chemistry; Plant Proteins - genetics; Plant Proteins - metabolism; Plant Structures - metabolism; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Salicylates - chemistry; Salicylates - metabolism; Sequence Homology, Amino Acid; Substrate Specificity - physiology; Tomatoes; Uridine Diphosphate Glucose - metabolism; Volatile Organic Compounds - metabolism
• ISSN: 1742-464X; 1742-4658
• DOI: 10.1111/j.1742-4658.2010.07962.x

The volatile compounds that constitute the fruit aroma of ripe tomato (Solanum lycopersicum) are often sequestered in glycosylated form. A homology-based screen was used to identify the gene SlUGT5, which is a member of UDP-glycosyltransferase 72 family and shows specificity towards a range of substrates, including flavonoid, flavanols, hydroquinone, xenobiotics and chlorinated pollutants. SlUGT5 was shown to be expressed primarily in ripening fruit and flowers, and mapped to chromosome I in a region containing a QTL that affected the content of guaiacol and eugenol in tomato crosses. Recombinant SlUGT5 protein demonstrated significant activity towards guaiacol and eugenol, as well as benzyl alcohol and methyl salicylate; however, the highest in vitro activity and affinity was shown for hydroquinone and salicyl alcohol. NMR analysis identified isosalicin as the only product of salicyl alcohol glycosylation. Protein modelling and substrate docking analysis were used to assess the basis for the substrate specificity of SlUGT5. The analysis correctly predicted the interactions with SlUGT5 substrates, and also indicated that increased hydrogen bonding, due to the presence of a second hydrophilic group in methyl salicylate, guaiacol and hydroquinone, appeared to more favourably anchor these acceptors within the glycosylation site, leading to increased stability, higher activities and higher substrate affinities.