Characterization and Engineering of Two Highly Paralogous Sesquiterpene Synthases Reveal a Regioselective Reprotonation Switch
© 2024 Wiley-VCH GmbH..
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.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:63 |
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| Enthalten in: |
Angewandte Chemie (International ed. in English) - 63(2024), 13 vom: 22. März, Seite e202315674 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Ye, Dan [VerfasserIn] |
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| Links: |
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| Themen: |
Alkyl and Aryl Transferases |
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| Anmerkungen: |
Date Completed 19.03.2024 Date Revised 19.03.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1002/anie.202315674 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM368162427 |
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| 245 | 1 | 0 | |a Characterization and Engineering of Two Highly Paralogous Sesquiterpene Synthases Reveal a Regioselective Reprotonation Switch |
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| 520 | |a 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 | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a functional interconversion | |
| 650 | 4 | |a germacrene A | |
| 650 | 4 | |a molecular basis | |
| 650 | 4 | |a regioselective reprotonation | |
| 650 | 4 | |a sesquiterpene synthase | |
| 650 | 7 | |a cyperene |2 NLM | |
| 650 | 7 | |a Sesquiterpenes |2 NLM | |
| 650 | 7 | |a Alkyl and Aryl Transferases |2 NLM | |
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| 700 | 1 | |a Shao, Yi-Zhen |e verfasserin |4 aut | |
| 700 | 1 | |a Li, Wen-Rui |e verfasserin |4 aut | |
| 700 | 1 | |a Cui, Zhen-Jia |e verfasserin |4 aut | |
| 700 | 1 | |a Gong, Ting |e verfasserin |4 aut | |
| 700 | 1 | |a Yang, Jin-Ling |e verfasserin |4 aut | |
| 700 | 1 | |a Wang, Hai-Qiang |e verfasserin |4 aut | |
| 700 | 1 | |a Dai, Jun-Gui |e verfasserin |4 aut | |
| 700 | 1 | |a Feng, Ke-Ping |e verfasserin |4 aut | |
| 700 | 1 | |a Ma, Ming |e verfasserin |4 aut | |
| 700 | 1 | |a Ma, Shuang-Gang |e verfasserin |4 aut | |
| 700 | 1 | |a Liu, Yun-Bao |e verfasserin |4 aut | |
| 700 | 1 | |a Zhu, Ping |e verfasserin |4 aut | |
| 700 | 1 | |a Yu, Shi-Shan |e verfasserin |4 aut | |
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