Structure and Mechanism of the Monoterpene Cyclolavandulyl Diphosphate Synthase that Catalyzes Consecutive Condensation and Cyclization

• Published in: Angewandte Chemie International Edition Vol. 56; no. 47; pp. 14913 - 14917
• Main Authors: Tomita, Takeo, Kobayashi, Masaya, Karita, Yuma, Yasuno, Yoko, Shinada, Tetsuro, Nishiyama, Makoto, Kuzuyama, Tomohisa
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 20.11.2017
• Edition: International ed. in English
• Subjects: biosynthesis; Chemical reactions; Condensation; crystal structure; cyclization; head-to-middle condensation; Magnesium; Mutagenesis; Terpene synthase; terpene synthases; crystal structure; head-to-middle condensation; terpene synthases; cyclization; biosynthesis
• ISSN: 1433-7851; 1521-3773
• DOI: 10.1002/anie.201708474

We report the three‐dimensional structure of cyclolavandulyl diphosphate (CLPP) synthase (CLDS), which consecutively catalyzes the condensation of two molecules of dimethylallyl diphosphate (DMAPP) followed by cyclization to form a cyclic monoterpene, CLPP. The structures of apo‐CLDS and CLDS in complex with Tris, pyrophosphate, and Mg2+ ion were refined at 2.00 Å resolution and 1.73 Å resolution, respectively. CLDS adopts a typical fold for cis‐prenyl synthases and forms a homo‐dimeric structure. An in vitro reaction using a regiospecifically 2H‐substituted DMAPP substrate revealed the intramolecular proton transfer mechanism of the CLDS reaction. The CLDS structure and structure‐based mutagenesis provide mechanistic insights into this unprecedented terpene synthase. The combination of structural and mechanistic insights advances the knowledge of intricate terpene synthase‐catalyzed reactions. A synthase that can do both: Cyclolavandulyl diphosphate (CLPP) synthase (CLDS) catalyzes “head‐to‐middle” condensation of two molecules of dimethylallyl diphosphate (DMAPP) and subsequent cyclization to produce CLPP. The crystal structure of CLDS in complex with Tris, pyrophosphate, and Mg2+ ion was refined at 1.73 Å resolution. An in vitro reaction using a regiospecifically deuterated DMAPP, the CLDS structure, and mutagenesis provide mechanistic insights into this unique terpene synthase.