Multi-substrate flavonol O-glucosyltransferases from strawberry (Fragaria×ananassa) achene and receptacle

In an effort to characterize fruit ripening-related genes functionally, two glucosyltransferases, FaGT6 and FaGT7, were cloned from a strawberry (Fragaria×ananassa) cDNA library and the full-length open reading frames were amplified by rapid amplification of cDNA ends. FaGT6 and FaGT7 were expressed heterologously as fusion proteins in Escherichia coli and target protein was purified using affinity chromatography. Both recombinant enzymes exhibited a broad substrate tolerance in vitro, accepting numerous flavonoids, hydroxycoumarins, and naphthols. FaGT6 formed 3-O-glucosides and minor amounts of 7-O-, 4′-O-, and 3′-O-monoglucosides and one diglucoside from flavonols such as quercetin. FaGT7 converted quercetin to the 3-O-glucoside and 4′-O-glucoside and minor levels of the 7- and 3′-isomers but formed no diglucoside. Gene expression studies showed that both genes are strongly expressed in achenes of small-sized green fruits, while the expression levels were generally lower in the receptacle. Significant levels of quercetin 3-O-, 7-O-, and 4′-O-glucosides, kaempferol 3-O- and 7-O-glucosides, as well as isorhamnetin 7-O-glucoside, were identified in achenes and the receptacle. In the receptacle, the expression of both genes is negatively controlled by auxin which correlates with the ripening-related gene expression in this tissue. Salicylic acid, a known signal molecule in plant defence, induces the expression of both genes. Thus, it appears that FaGT6 and FaGT7 are involved in the glucosylation of flavonols and may also participate in xenobiotic metabolism. The latter function is supported by the proven ability of strawberries to glucosylate selected unnatural substrates injected in ripe fruits. This report presents the first biochemical characterization of enzymes mainly expressed in strawberry achenes and provides the foundation of flavonoid metabolism in the seeds

Phylogenetic analysis of selected plant secondary product glycosyltransferases

The neighbor-joining tree was calculated with the Treecon software package (). Distance calculation was performed with Poisson correction and insertions/deletions were not taken into account. The tree was rooted using a sterol glycosyltransferase from (AsSGT) as an outgroup. Branch lengths indicate the number of substitutions per site. Bootstrap analysis was performed with 100 replicates and only values above 50% are shown. GenBank accession numbers and sources for the respective protein sequences are: AtUGT73B4, AAD17393 (); AtUGT73B5, AAD17392 (); AtUGT73B2, AAR01231 (); DicGT4, BAD52006 (); DbBet5GT, CAB56231 (); SbF7G, BAA83484 (); Letwi1, CAA59450 (); NtIS10a, AAB36652 (); NtIS5a, AAB36653 (); FaGT7, ABB 92749 (×); AtUGT71C2, AAC35238 (); AtUGT71D1, AAC35239 (); DicGT2, BAD52004 (); DbBet6GT, AAL57240 (); AtUGT71B6, BAB02840 (); FaGT6, ABB92748 (×); FaGT3, AAU09444 (×); NtGT3, BAB88934 (); NtGT1b, BAB60721 (); NtGT1a, BAB60720 (); NtSalGT, AAF61647 (); AtUGT84B1, AAB87119 (); AtUGT84B2, AAB87106 (); CuLimGT, BAA93039 (); FaGT2, AAU09443 (×); FaGT5, ABB92747 (); AtUGT84A3, CAB10327 (); AtUGT84A1, CAB10326 (); AtUGT84A2, BAB02351 (); BnSinGT, AAF98390 (); FaGT4, AAU09445 (×); PhA3RhaT, CAA81057 (); Ip3GGT, BAD95882 (); In3GGT, BAD95881 (); GtF3GT, BAA12737 (); DicGT3, BAD52005 (); VmUFGT1, BAA36972 (); PhF3GalT, AAD55985 (); DicGT1, BAD52003 (); VvGT1, AAB81682 (); FaGT1, AAU09442 (×); AtUGT78D1, NP_564357 (); AtUGT78D2, NP_197207 (); AsSGT, CAB06081 ().<p><b>Copyright information:</b></p><p>Taken from "Multi-substrate flavonol -glucosyltransferases from strawberry () achene and receptacle"</p><p></p><p>Journal of Experimental Botany 2008;59(10):2611-2625.</p><p>Published online 17 May 2008</p><p>PMCID:PMC2486459.</p><p></p

Sequence alignment of quercetin and kaempferol -glucosyltransferases, including two already crystallized proteins from (VvGT1) and (UGT71G1) as well as × GT6 (FaGT6), and × GT7 (FaGT7)

The alignment was performed using ClustalX (). Conserved amino acids are shaded, amino acids within 5 Å to kaempferol in the protein model of VvGT are boxed.<p><b>Copyright information:</b></p><p>Taken from "Multi-substrate flavonol -glucosyltransferases from strawberry () achene and receptacle"</p><p></p><p>Journal of Experimental Botany 2008;59(10):2611-2625.</p><p>Published online 17 May 2008</p><p>PMCID:PMC2486459.</p><p></p

Gene expression of (A) and (B) in achenes (black columns) and the receptacle (open columns) of small-sized green (G) and red (R) fruits

Transcript levels were analysed by qPCR as described in Materials and methods. Gene expression is shown as relative expression normalized with receptacle tissue from small-sized green fruit (G). Hormonal control of (C) and (D) gene expression. The achenes were carefully removed at mid-sized green stage and the fruits were harvested after 5 d. Additionally, deachened green fruits were treated with a lanolin paste containing the synthetic auxin NAA. Gene expression was analysed by qPCR as described in Materials and methods and the data were normalized against untreated strawberries with the achenes still attached to the fruit.<p><b>Copyright information:</b></p><p>Taken from "Multi-substrate flavonol -glucosyltransferases from strawberry () achene and receptacle"</p><p></p><p>Journal of Experimental Botany 2008;59(10):2611-2625.</p><p>Published online 17 May 2008</p><p>PMCID:PMC2486459.</p><p></p

On the Visibility of Al Surface Sites of γ-Alumina: A Combined Computational and Experimental Point of View

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