title: 4-Coumaroyl coenzyme A 3-hydroxylase activity from cell cultures of Lithospermum erythrorhizon and its relationship to polyphenol oxidase.
abstract: A 4-coumaroyl-CoA 3-hydroxylase activity was purified 4600-fold from cell cultures of Lithospermum erythrorhizon. The enzyme showed a molecular mass of 42,400 +/- 1700 Da in gel chromatography and required ascorbate, NADH, or NADPH as cofactors. 4-Coumaroyl-CoA, 4-coumarate, p-cresol, and several other phenolic substances, but not tyrosine, were accepted as substrates for the hydroxylation. Besides hydroxylase activity, the enzyme showed diphenol oxidase activity. Both activities were inhibited by diethyldithiocarbamate or beta-mercaptoethanol, although at different concentrations. The enzyme showed striking similarity to a 4-coumaroyl-glucose 3-hydroxylase from sweet potato (Ipomoe batatas) roots, which has reportedly been purified to homogeneity and identified as a specific enzyme of chlorogenic acid biosynthesis. Close examination and comparison to a commercially available polyphenol oxidase, however, suggest that the enzyme activities purified from both Lithospermum and sweet potato are polyphenol oxidases rather than specific enzymes of secondary metabolism.
authors.affiliation: Pharmazeutisches Institut, Eberhard-Karls Universität Tübingen, Auf der Morgenstelle 8, Tübingen, 72076, Germany.
authors.forename: Z X
authors.initials: ZX
authors.lastname: Wang
authors.forename: S M
authors.initials: SM
authors.lastname: Li
authors.forename: R
authors.initials: R
authors.lastname: Löscher
authors.forename: L
authors.initials: L
authors.lastname: Heide
mesh_terms: D000227:Adenine Nucleotides / Q000378:metabolism; D001205:Ascorbic Acid / Q000378:metabolism; D004156:Catechol Oxidase / Q000378:metabolism*; D002478:Cells, Cultured; D004579:Electron Transport; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008670:Metals / Q000494:pharmacology; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D010084:Oxidation-Reduction; D010940:Plant Proteins / Q000378:metabolism; D010944:Plants / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D000227:Adenine Nucleotides; D004791:Enzyme Inhibitors; D008670:Metals; D010940:Plant Proteins; D006899:Mixed Function Oxygenases; D004156:Catechol Oxidase; C066923:4-coumaroyl-D-glucose hydroxylase; C058643:4-coumaroyl-coenzyme A 3-hydroxylase; D001205:Ascorbic Acid