倍半萜
区域选择性
脱质子化
质子化
ATP合酶
化学
立体化学
酶
生物化学
有机化学
催化作用
离子
作者
Dan Ye,Yizhen Shao,Wenrui Li,Zhen Cui,Ting Gong,Jin-Ling Yang,Hai-Qiang Wang,Jungui Dai,Ke-Ping Feng,Ming Ma,Shuang‐Gang Ma,Yun‐Bao Liu,Ping Zhu,Shi‐Shan Yu
标识
DOI:10.1002/anie.202315674
摘要
Abstract Sesquiterpene synthases (STPSs) catalyze carbocation‐driven cyclization reactions that can generate structurally diverse hydrocarbons. The deprotonation‐reprotonation process is widely used in STPSs to promote structural diversity, largely attributable to the distinct regio/stereoselective reprotonations. However, the molecular basis for reprotonation regioselectivity remains largely understudied. Herein, we analyzed two highly paralogous STPSs, Artabotrys hexapetalus (−)‐cyperene synthase (AhCS) and ishwarane synthase (AhIS), which catalyze reactions that are distinct from the regioselective protonation of germacrene A (GA), resulting in distinct skeletons of 5/5/6 tricyclic (−)‐cyperene and 6/6/5/3 tetracyclic ishwarane, respectively. Isotopic labeling experiments demonstrated that these protonations occur at C3 and C6 of GA in AhCS and AhIS, respectively. The cryo‐electron microscopy‐derived AhCS complex structure provided the structural basis for identifying different key active site residues that may govern their functional disparity. The structure‐guided mutagenesis of these residues resulted in successful functional interconversion between AhCS and AhIS, thus targeting the three active site residues [L311‐S419‐C458]/[M311‐V419‐A458] that may act as a C3/C6 reprotonation switch for GA. These findings facilitate the rational design or directed evolution of STPSs with structurally diverse skeletons.
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