title: Novel flavonol 3-sulfotransferase. Purification, kinetic properties, and partial amino acid sequence.
abstract: A flavonol sulfotransferase (EC 2.8.2.-), which catalyzes the transfer of the sulfate group from 3'-phosphoadenosine 5'-phosphosulfate to the 3-hydroxyl group of flavonol aglycones, has been purified to apparent homogeneity from Flaveria chloraefolia. The specific activity of flavonol 3-sulfotransferase was enriched 2000-fold, as compared with the homogenate, with a recovery of 9%. The molecular mass of the native and denatured enzyme was found to be 34.5 kDa, suggesting that the active from of the enzyme is a monomer. The enzyme exhibited expressed specificity for position 3 of flavonol aglycones, showed two activity optima at pH 6.0 and 8.5, did not require divalent cations, and was not inhibited by either EDTA or sulfhydryl group reagents. The results of substrate interaction kinetics and product inhibition are consistent with an Ordered Bi Bi mechanism where 3'-phosphoadenosine 5'-phosphosulfate is the first substrate to bind to the enzyme and 3'-phosphoadenosine 5'-phosphate is the final product to be released. The amino acid sequence of two peptides representing 17 and 33 amino acids showed no significant sequence similarity with the amino acid sequences reported for animal sulfotransferases. Antibodies raised against F. chloraefolia 3-sulfotransferase were found to cross-react with the 3'- and 4'-sulfotransferase activities of the same plant, suggesting that the three enzymes are structurally related.
authors.affiliation: Department of Biology, Concordia University, Montreal, Quebec, Canada.
authors.forename: L
authors.initials: L
authors.lastname: Varin
authors.forename: R K
authors.initials: RK
authors.lastname: Ibrahim
mesh_terms: D000595:Amino Acid Sequence; D002845:Chromatography; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D017886:Durapatite; D004591:Electrophoresis, Polyacrylamide Gel; D006882:Hydroxyapatites; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D010944:Plants / Q000201:enzymology*; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D015238:Sulfotransferases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D006882:Hydroxyapatites; D010446:Peptide Fragments; D017886:Durapatite; C022321:PAPS sulfotransferase; D015238:Sulfotransferases