title: Cyclization mechanism of phomopsene synthase: mass spectrometry based analysis of various site-specifically labeled terpenes.
abstract: Elucidation of the cyclization mechanism catalyzed by terpene synthases is important for the rational engineering of terpene cyclases. We developed a chemoenzymatic method for the synthesis of systematically deuterium-labeled geranylgeranyl diphosphate (GGPP), starting from site-specifically deuterium-labeled isopentenyl diphosphates (IPPs) using IPP isomerase and three prenyltransferases. We examined the cyclization mechanism of tetracyclic diterpene phomopsene with phomopsene synthase. A detailed EI-MS analysis of phomopsene labeled at various positions allowed us to propose the structures corresponding to the most intense peaks, and thus elucidate a cyclization mechanism involving double 1,2-alkyl shifts and a 1,2-hydride shift via a dolabelladien-15-yl cation. Our study demonstrated that this newly developed method is highly sensitive and provides sufficient information for a reliable assignment of the structures of fragmented ions.
authors.affiliation: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Sandip S
authors.initials: SS
authors.lastname: Shinde
authors.affiliation: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Minami
authors.affiliation: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Tetsuo
authors.initials: T
authors.lastname: Tokiwano
authors.affiliation: Department of Bioresource Engineering, Yamagata University, Yamagata, Japan.
authors.forename: Tomonobu
authors.initials: T
authors.lastname: Toyomasu
authors.affiliation: The Institute of Scientific and Industrial Research, Osaka University, Osaka, Japan.
authors.forename: Nobuo
authors.initials: N
authors.lastname: Kato
authors.affiliation: Department of Bioresource Engineering, Yamagata University, Yamagata, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sassa
authors.affiliation: Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Oikawa
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000378:metabolism*; D003500:Cyclization; D003903:Deuterium / Q000737:chemistry; D045782:Hemiterpenes / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D009943:Organophosphorus Compounds / Q000737:chemistry; D011106:Polyisoprenyl Phosphates / Q000138:chemical synthesis*; D013729:Terpenes / Q000737:chemistry*
chemical_list: D045782:Hemiterpenes; D009943:Organophosphorus Compounds; D011106:Polyisoprenyl Phosphates; D013729:Terpenes; C004809:isopentenyl pyrophosphate; D003903:Deuterium; D019883:Alkyl and Aryl Transferases; C108877:terpene synthase; C002963:geranylgeranyl pyrophosphate