title: Predicting the substrate specificity of a glycosyltransferase implicated in the production of phenolic volatiles in tomato fruit.
abstract: 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.
authors.affiliation: Université de Toulouse, UMR Génomique et Biotechnologie des Fruits, INRA-INP/ENSAT, Castanet-Tolosan, France.
authors.forename: Thomas
authors.initials: T
authors.lastname: Louveau
authors.forename: Celine
authors.initials: C
authors.lastname: Leitao
authors.forename: Sol
authors.initials: S
authors.lastname: Green
authors.forename: Cyril
authors.initials: C
authors.lastname: Hamiaux
authors.forename: Benoît
authors.initials: B
authors.lastname: van der Rest
authors.forename: Odile
authors.initials: O
authors.lastname: Dechy-Cabaret
authors.forename: Ross G
authors.initials: RG
authors.lastname: Atkinson
authors.forename: Christian
authors.initials: C
authors.lastname: Chervin
mesh_terms: D000595:Amino Acid Sequence; D019905:Benzyl Alcohol / Q000737:chemistry / Q000378:metabolism; D001592:Benzyl Alcohols / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain / Q000502:physiology; D002875:Chromosomes / Q000235:genetics; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005054:Eugenol / Q000737:chemistry / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics; D016695:Glycosyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006139:Guaiacol / Q000737:chemistry / Q000378:metabolism; D006860:Hydrogen Bonding; D006873:Hydroquinones / Q000737:chemistry / Q000378:metabolism; D018551:Solanum lycopersicum / Q000201:enzymology* / Q000235:genetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010636:Phenols / Q000378:metabolism*; D010626:Phenylethyl Alcohol / Q000737:chemistry / Q000378:metabolism; D010802:Phylogeny; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018514:Plant Structures / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012459:Salicylates / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity / Q000502:physiology; D014532:Uridine Diphosphate Glucose / Q000378:metabolism; D055549:Volatile Organic Compounds / Q000378:metabolism*
chemical_list: D001592:Benzyl Alcohols; D006873:Hydroquinones; D010636:Phenols; D010940:Plant Proteins; D011994:Recombinant Proteins; D012459:Salicylates; D055549:Volatile Organic Compounds; C018966:4-hydroxybenzyl alcohol; D005054:Eugenol; D006139:Guaiacol; D016695:Glycosyltransferases; C010631:salicyl alcohol; C033069:methyl salicylate; D019905:Benzyl Alcohol; D010626:Phenylethyl Alcohol; D014532:Uridine Diphosphate Glucose; C031927:hydroquinone