title: Purification of an UDP-glucose:flavone, 7-O-glucosyltransferase, from Silene latifolia using a specific interaction between the enzyme and phenyl-Sepharose.
abstract: An UDP-glucose:flavonoid, 7-O-glucosyltransferase, from Silene latifolia was isolated from petals and purified 450-fold using a combination of gel-filtration, affinity chromatography and anion-exchange chromatography. Affinity chromatography on a phenyl-Sepharose CL-4B column in combination with elution with the substrate, isovitexin (6-C-glucosylapigenin), was an especially effective purification step. A purification factor between 10 and 20 could be reached using this column. A possible mechanism for the specific interaction of the enzyme with the phenyl-Sepharose will be discussed. This method of purification may also be applicable to other enzymes which use aromatic compounds as substrates. On a SDS-PAGE gel a band of 54 kDa, which co-purified with enzyme activity, could be detected in the purest fraction.
authors.affiliation: University of Utrecht, Department of Plant Ecology and Evolutionary Biology, The Netherlands.
authors.forename: P
authors.initials: P
authors.lastname: Vellekoop
authors.forename: L
authors.initials: L
authors.lastname: Lugones
authors.forename: J
authors.initials: J
authors.lastname: van Brederode
mesh_terms: D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005964:Glucosyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D010944:Plants / Q000201:enzymology*; D012685:Sepharose / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: C020927:phenyl-sepharose; D012685:Sepharose; D005964:Glucosyltransferases; C083090:flavone 7-O-beta-glucosyltransferase