Jiadian Wang1,2,3, Ping Su4, Linhui Gao5, Yifeng Zhang1, Jian Wang3, Lichan Tu1, Yujun Zhao3, Yun Lu1, Yan Yin6, Luqi Huang3, Wei Gao7,8. 1. Department of Traditional Chinese Medicine, Capital Medical University, Beijing, 100069, China. 2. Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China. 3. National Resource Center for Chinese Materia Medica, Chinese Academy of Chinese Medical Sciences, Beijing, 100700, China. 4. Department of Chemistry, The Scripps Research Institute, Jupiter, FL, 33458, USA. 5. Division of Biotechnology, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China. 6. School of Chinese Pharmacy, Beijing University of Chinese Medicine, Beijing, 102488, China. 7. Department of Traditional Chinese Medicine, Capital Medical University, Beijing, 100069, China. weigao@ccmu.edu.cn. 8. Beijing Shijitan Hospital, Capital Medical University, Beijing, 100038, China. weigao@ccmu.edu.cn.
Abstract
MAIN CONCLUSION: A novel cytochrome P450 from Tripterygium wilfordii, CYP81AM1, specifically catalyses the C-15 hydroxylation of dehydroabietic acid. This is the first CYP450 to be found in plants with this function. Cytochrome P450 oxygenases (CYPs) play an important role in the post-modification in biosynthesis of plant bioactive terpenoids. Here, we found that CYP81AM1 can catalyze the formation of 15-hydroxydehydroabietic acid by in vitro enzymatic reactions and in vivo yeast feeding assays. This is the first study to show that CYP81 family enzymes are involved in the hydroxylation of abietane diterpenoids. At the same time, we found that CYP81AM1 could not catalyse abietatriene and dehydroabietinol, suggesting that the occurrence of the reaction may be related to the carboxyl group. Through molecular docking and site mutations, it was found that some amino acid sites (F104, K107) near the carboxyl group had an important effect on enzyme activity, also suggesting that the carboxyl group played an important role in the occurrence of the reaction.
MAIN CONCLUSION: A novel cytochrome P450 from Tripterygium wilfordii, CYP81AM1, specifically catalyses the C-15 hydroxylation of dehydroabietic acid. This is the first CYP450 to be found in plants with this function. Cytochrome P450 oxygenases (CYPs) play an important role in the post-modification in biosynthesis of plant bioactive terpenoids. Here, we found that CYP81AM1 can catalyze the formation of 15-hydroxydehydroabietic acid by in vitro enzymatic reactions and in vivo yeast feeding assays. This is the first study to show that CYP81 family enzymes are involved in the hydroxylation of abietane diterpenoids. At the same time, we found that CYP81AM1 could not catalyse abietatriene and dehydroabietinol, suggesting that the occurrence of the reaction may be related to the carboxyl group. Through molecular docking and site mutations, it was found that some amino acid sites (F104, K107) near the carboxyl group had an important effect on enzyme activity, also suggesting that the carboxyl group played an important role in the occurrence of the reaction.
Authors: Allison M Colthart; Drew R Tietz; Yuhua Ni; Jessica L Friedman; Marina Dang; Thomas C Pochapsky Journal: Sci Rep Date: 2016-02-25 Impact factor: 4.379