| Abstract Detail
Secondary Metabolism Blount, J. W. [1], Modolo, L. V. [1], Achnine, L. [2], Dixon, R. A. [1]. Expression and in vitro Characterization of Eight Medicago truncatula (Iso)flavonoid Glycosyltransferases. Glycosyltransferases (GTs) aid in the stabilization, compartmentation and activation of a variety of secondary metabolites by transferring nucleotide-diphosphate-activated sugars to low molecular weight compounds. The model legume Medicago truncatula contains many glycosylated secondary metabolites such as isoflavonoids, flavonoids, anthocyanins, and triterpene saponins, several of which are known have estrogenic, anti-inflammatory, and/or anticancer activity. An efficient HPLC method was developed to analyze candidate GTs against 32 potential substrates by dividing the substrates “equally” into four groups. The compounds in each group are readily separated by the HPLC method, and each compound within a particular group has a distinct UV spectrum. Using our extensive M. truncatula EST database, we have identified 63 full-length GTs which may be involved in the glycosylation of secondary metabolites. Soluble recombinant protein was purified for eighteen of the GTs using the Magne-His protein purification kit and assayed for activity using the four substrate mixtures and either UDP-glucose or UDP-glucuronic acid as the sugar donor. Eight of the eighteen GTs could glycosylate one or more of the (iso)flavonoid compounds in the mixtures, preferring UDP-glucose over UDP-glucuronic acid as the sugar donor. These eight GTs were further assayed for activity using UDP-glucose and a variety of individual flavonoid substrates including chalcones, coumestans, flavanones, flavones, flavonols, isoflavones, and pterocarpans. This poster will examine the in vitro glycosylation patterns of these eight GTs with the individual flavonoid substrates, identifying several of the preferred glycosylation sites.
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1 - The Samuel Roberts Noble Foundation, Plant Biology Division, 2510 Sam Noble Parkway, Ardmore, OK, 73401, USA
2 - GenApps, Inc., 4274 Colby Road, Winchester, Kentucky, 40391, USA
Keywords:
glycosyltransferase
Medicago truncatula
HPLC
enzyme assays
UDP-glucose
isoflavonoid
flavonoid.
Presentation Type: Plant Biology Abstract
Session: P
Location: Exhibit Hall (Northeast, Southwest & Southeast)/Hilton
Date: Sunday, July 8th, 2007
Time: 8:00 AM
Number: P20007
Abstract ID:175 |
