title: Identification and characterization of a GDSL lipase-like protein that catalyzes the ester-forming reaction for pyrethrin biosynthesis in Tanacetum cinerariifolium- a new target for plant protection.
abstract: Although natural insecticides pyrethrins produced by Tanacetum cinerariifolium are used worldwide to control insect pest species, little information is known of their biosynthesis. From the buds of T. cinerariifolium, we have purified a protein that is able to transfer the chrysanthemoyl group from the coenzyme A (CoA) thioester to pyrethrolone to produce pyrethrin I and have isolated cDNAs that encode the enzyme. To our surprise, the active principle was not a member of a known acyltransferase family but a member of the GDSL lipase family. The recombinant enzyme (TcGLIP) was expressed in Escherichia coli and displayed the acyltransferase reaction with high substrate specificity, recognized the absolute configurations of three asymmetric carbons and also showed esterase activity. A S40A mutation in the Block I domain reduced both acyltransferase and esterase activities, which suggested an important role of this serine residue in these two activities. The signal peptide directed the localization of TcGLIP::enhanced green fluorescent protein (EGFP) fusion, as well as EGFP, to the extracellular space. High TcGLIP gene expression was observed in the leaves of mature plants and seedlings as well as in buds and flowers, a finding that was consistent with the pyrethrin I content in these parts. Expression was enhanced in response to wounding, which suggested that the enzyme plays a key role in the defense mechanism of T. cinerariifolium.
authors.affiliation: Dainihon Jochugiku Co., Ltd., 1-1-11 Daikoku-cho, Toyonaka, Osaka 561-0827, Japan.
authors.forename: Yukio
authors.initials: Y
authors.lastname: Kikuta
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Ueda
authors.forename: Masafumi
authors.initials: M
authors.lastname: Takahashi
authors.forename: Tomonori
authors.initials: T
authors.lastname: Mitsumori
authors.forename: Gen
authors.initials: G
authors.lastname: Yamada
authors.forename: Koji
authors.initials: K
authors.lastname: Sakamori
authors.forename: Kengo
authors.initials: K
authors.lastname: Takeda
authors.forename: Shogo
authors.initials: S
authors.lastname: Furutani
authors.forename: Koji
authors.initials: K
authors.lastname: Nakayama
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Katsuda
authors.forename: Akikazu
authors.initials: A
authors.lastname: Hatanaka
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Matsuda
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D011723:Chrysanthemum cinerariifolium / Q000737:chemistry / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D004950:Esterases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004952:Esters; D035264:Flowers / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression / Q000235:genetics; D007306:Insecticides / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008049:Lipase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018515:Plant Leaves / Q000201:enzymology / Q000235:genetics; D018517:Plant Roots / Q000201:enzymology / Q000235:genetics; D018547:Plant Stems / Q000201:enzymology / Q000235:genetics; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D011722:Pyrethrins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D036226:Seedlings / Q000201:enzymology / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D004952:Esters; D007306:Insecticides; D021382:Protein Sorting Signals; D011722:Pyrethrins; D011993:Recombinant Fusion Proteins; C007051:chrysanthemic acid; D000217:Acyltransferases; D004950:Esterases; D008049:Lipase; C009119:pyrethrin I