Biosynthesis of the microtubule-destabilizing diterpene pseudolaric acid B from golden larch involves an unusual diterpene synthase.

Autor:	Mafu S; Department of Plant Biology, University of California, Davis, CA 95616., Karunanithi PS; Department of Plant Biology, University of California, Davis, CA 95616., Palazzo TA; Department of Chemistry, University of California, Davis, CA 95616., Harrod BL; Department of Chemistry, University of California, Davis, CA 95616., Rodriguez SM; Department of Plant Biology, University of California, Davis, CA 95616., Mollhoff IN; Department of Plant Biology, University of California, Davis, CA 95616., O'Brien TE; Department of Chemistry, University of California, Davis, CA 95616., Tong S; West Coast Metabolomics Center, University of California, Davis, CA 95616., Fiehn O; West Coast Metabolomics Center, University of California, Davis, CA 95616.; Biochemistry Department, King Abdulaziz University, Jeddah 23218, Saudi Arabia., Tantillo DJ; Department of Chemistry, University of California, Davis, CA 95616., Bohlmann J; Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T 1Z4, Canada., Zerbe P; Department of Plant Biology, University of California, Davis, CA 95616; pzerbe@ucdavis.edu.
Jazyk:	angličtina
Zdroj:	Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2017 Jan 31; Vol. 114 (5), pp. 974-979. Date of Electronic Publication: 2017 Jan 17.
DOI:	10.1073/pnas.1612901114
Abstrakt:	The diversity of small molecules formed via plant diterpene metabolism offers a rich source of known and potentially new biopharmaceuticals. Among these, the microtubule-destabilizing activity of pseudolaric acid B (PAB) holds promise for new anticancer agents. PAB is found, perhaps uniquely, in the coniferous tree golden larch (Pseudolarix amabilis, Pxa). Here we describe the discovery and mechanistic analysis of golden larch terpene synthase 8 (PxaTPS8), an unusual diterpene synthase (diTPS) that catalyzes the first committed step in PAB biosynthesis. Mining of the golden larch root transcriptome revealed a large TPS family, including the monofunctional class I diTPS PxaTPS8, which converts geranylgeranyl diphosphate into a previously unknown 5,7-fused bicyclic diterpene, coined "pseudolaratriene." Combined NMR and quantum chemical analysis verified the structure of pseudolaratriene, and co-occurrence with PxaTPS8 and PAB in P amabilis tissues supports the intermediacy of pseudolaratriene in PAB metabolism. Although PxaTPS8 adopts the typical three-domain structure of diTPSs, sequence phylogeny places the enzyme with two-domain TPSs of mono- and sesqui-terpene biosynthesis. Site-directed mutagenesis of PxaTPS8 revealed several catalytic residues that, together with quantum chemical calculations, suggested a substantial divergence of PxaTPS8 from other TPSs leading to a distinct carbocation-driven reaction mechanism en route to the 5,7-trans-fused bicyclic pseudolaratriene scaffold. PxaTPS8 expression in microbial and plant hosts provided proof of concept for metabolic engineering of pseudolaratriene. Competing Interests: The authors declare no conflict of interest.
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu