Kekulé-Institut für Organische Chemie und Biochemie, Rheinische Friedrich-Wilhelms-Universität Bonn, Gerhard-Domagk-Straße 1, 53121, Bonn, Germany.
Institut für Chemie und Biochemie - Organische Chemie, Freie Universität Berlin, Takustraße 3, 14195, Berlin, Germany.
Angew Chem Int Ed Engl. 2016 Jul 18;55(30):8748-51. doi: 10.1002/anie.201603782. Epub 2016 Jun 13.
Two sesquiterpene cyclases from Fusarium fujikuroi were expressed in Escherichia coli and purified. The first enzyme was inactive because of a critical mutation, but activity was restored by sequence correction through site-directed mutagenesis. The mutated enzyme and two naturally functional homologues from other fusaria converted farnesyl diphosphate into guaia-6,10(14)-diene. The second enzyme produced eremophilene. The absolute configuration of guaia-6,10(14)-diene was elucidated by enantioselective synthesis, while that of eremophilene was evident from the sign of its optical rotation and is opposite to that in plants but the same as in Sorangium cellulosum. The mechanisms of both terpene cyclases were studied with various (13) C- and (2) H-labelled FPP isotopomers.
两株来自藤仓镰刀菌的倍半萜环化酶在大肠杆菌中表达并纯化。第一个酶由于关键突变而失活，但通过定点突变进行序列校正恢复了活性。突变酶和来自其他真菌的两种天然功能同源物将法呢基二磷酸转化为愈创木-6,10(14)-二烯。第二种酶产生了埃罗莫芬。通过对映选择性合成阐明了愈创木-6,10(14)-二烯的绝对构型，而埃罗莫芬的构型则从其旋光性的符号明显看出，与植物中的相反，但与 Sorangium cellulosum 中的相同。用各种（13）C 和（2）H 标记的 FPP 同位素标记物研究了两种萜烯环化酶的机制。
