Triterpenoids are valuable medicinal scaffolds,characterized by excellent pharmacological properties and the presence of hydroxyl and carboxyl groups that allow for further structural modifications.Expanding the scope...Triterpenoids are valuable medicinal scaffolds,characterized by excellent pharmacological properties and the presence of hydroxyl and carboxyl groups that allow for further structural modifications.Expanding the scope of oxidative modifications on these molecules is crucial for increasing their synthetic structural diversity and unlocking new potential pharmacological activities.However,the progress has been limited by the scarcity of suitable tailoring enzymes.Here,we reported a break-through in achieving targeted and remote dual-site oxidation of licorice triterpenoids using a single P450 mutant.This approach successfully enabled the selective synthesis of the rare triterpenoid,liquiritic acid and 24-OH-liquiritic acid.Our findings demonstrate that microenvironmental accessibility engineering of triterpenoid substrates within the P450 enzyme is essential for continuous and regioselective oxidation.This study not only sheds light on the mechanistic aspects of P450 catalysis but also expands the enzymatic toolkit for selective oxidative modifications in triterpenoid biosynthesis.展开更多
Many labdane-related diterpenoids(LRDs)exhibit high values in drug development.Their diversity in structure and bioactivity,to a large extent,arise from oxidative modifications which are mainly catalyzed by cytochrome...Many labdane-related diterpenoids(LRDs)exhibit high values in drug development.Their diversity in structure and bioactivity,to a large extent,arise from oxidative modifications which are mainly catalyzed by cytochrome P450s(CYPs).The medicinal plant Euphorbia fischeriana Steud.is rich in LRDs with distinct scaffolds.Herein,we characterized three cytochrome P450s involved in LRD biosynthesis from this plant.Notably,CYP71D450 and CYP701A148 are two substrate-promiscuity CYPs.The former is the first example of CYPs which can oxidize C-3 of ent-atisane skeleton and ent-isopimara-7(8),15-diene,and the latter is the first example of CYPs which can oxidize C-19 of ent-abietane and ent-pimarane skeletons.This study expands the toolkit for bioproduction of diverse LRDs.展开更多
Toxic heavy metal and metalloid(THMM)contamination poses a major global challenge,threatening human health and sustainable agriculture.The crucial role of the Cytochrome P450(CYP)gene family in plant tolerance to THMM...Toxic heavy metal and metalloid(THMM)contamination poses a major global challenge,threatening human health and sustainable agriculture.The crucial role of the Cytochrome P450(CYP)gene family in plant tolerance to THMMs has been recently highlighted,but there is still a lack of comprehensive understanding,especially in relation to metabolites.This study delved into the identification of CYP genes that are linked to the tolerance mechanisms of plants in response to heavy metal stress.The findings highlight the significant metabolic pathways that contribute to this resilience,using rice and Arabidopsis as exemplars.THMM exposure changed CYP gene expression in plants,and THMM antidotes mitigated its downregulation and that of flavonoid biosynthesis genes.CYP genes involved in THMM responses were predominantly enriched in the pathways associated with flavonoid synthesis,indicating functional adaptations to distinct stresses.Notably,anthocyanin(Ant)accumulation,a type of flavonoid,affected the uptake of various heavy metals in Brassica rapa,with flavonoid biosynthesis-associated genes correlating with Cd or As tolerance and Ant content.These findings highlight the critical importance of flavonoid metabolism and the intricate network of biosynthesis genes in bolstering plant resilience against heavy metal stress.This enhanced understanding paves the way for significant advancements in phytoremediation technologies,offering innovative solutions for soil and water decontamination.展开更多
细胞色素P450酶是自然界最大的一类氧化还原酶超家族,广泛参与药物代谢、植物次生代谢以及环境污染物转化等重要生物过程。为了生成具有高保真度和多样性的人工P450酶序列,本研究提出了一种基于扩散模型的P450酶序列生成方法——P450Di...细胞色素P450酶是自然界最大的一类氧化还原酶超家族,广泛参与药物代谢、植物次生代谢以及环境污染物转化等重要生物过程。为了生成具有高保真度和多样性的人工P450酶序列,本研究提出了一种基于扩散模型的P450酶序列生成方法——P450Diff2。该模型基于参数量达6.4亿的EvoDiff-Seq框架进行构建,并以来自NCBI、GMind注释、RNA-Seq组装和宏基因组数据库的1041254条非冗余P450蛋白序列作为训练数据。基于训练完成的模型进行序列生成评估,结果显示,P450Diff2在氨基酸组成分布、序列特征空间、序列相似性分布以及结构合理性等多个维度均优于此前提出的P450Diffusion模型,其所生成序列的平均局部预测置信度(predicted local distance difference test,pLDDT)达到72.29。实验结果进一步表明,所生成的序列中有60%能够正确折叠为具有生物活性的P450酶,表明本方法不仅有效保留了天然序列的结构特征,同时具备一定的功能性序列生成可能性。结合大规模序列生成与筛选流程,有望快速设计生成高效新酶序列,降低实验筛选的时间与成本,为新酶的高效、可扩展设计提供了全新的参考。展开更多
基金supported by grants from the National Natural Science Foundation of China(Nos.22108154,22138006,32171430).
文摘Triterpenoids are valuable medicinal scaffolds,characterized by excellent pharmacological properties and the presence of hydroxyl and carboxyl groups that allow for further structural modifications.Expanding the scope of oxidative modifications on these molecules is crucial for increasing their synthetic structural diversity and unlocking new potential pharmacological activities.However,the progress has been limited by the scarcity of suitable tailoring enzymes.Here,we reported a break-through in achieving targeted and remote dual-site oxidation of licorice triterpenoids using a single P450 mutant.This approach successfully enabled the selective synthesis of the rare triterpenoid,liquiritic acid and 24-OH-liquiritic acid.Our findings demonstrate that microenvironmental accessibility engineering of triterpenoid substrates within the P450 enzyme is essential for continuous and regioselective oxidation.This study not only sheds light on the mechanistic aspects of P450 catalysis but also expands the enzymatic toolkit for selective oxidative modifications in triterpenoid biosynthesis.
基金supported by grants from the National Natural Science Foundation of China[Nos.82474024,82293682(82293680),82073953]the Nonprofit Central Research Institute Fund of Chinese Academy of Medical Sciences(No.2021-RC350-009)+1 种基金the CAMS Innovation Fund for Medical Sciences(No.2023-I2M-2-006)the Special Fund for the Construction of Qinghai Innovation Platform(No.2024-ZJ-T02)。
文摘Many labdane-related diterpenoids(LRDs)exhibit high values in drug development.Their diversity in structure and bioactivity,to a large extent,arise from oxidative modifications which are mainly catalyzed by cytochrome P450s(CYPs).The medicinal plant Euphorbia fischeriana Steud.is rich in LRDs with distinct scaffolds.Herein,we characterized three cytochrome P450s involved in LRD biosynthesis from this plant.Notably,CYP71D450 and CYP701A148 are two substrate-promiscuity CYPs.The former is the first example of CYPs which can oxidize C-3 of ent-atisane skeleton and ent-isopimara-7(8),15-diene,and the latter is the first example of CYPs which can oxidize C-19 of ent-abietane and ent-pimarane skeletons.This study expands the toolkit for bioproduction of diverse LRDs.
基金supported by the Guizhou Provincial Science and Technology Project(Grant No.[2022]091).
文摘Toxic heavy metal and metalloid(THMM)contamination poses a major global challenge,threatening human health and sustainable agriculture.The crucial role of the Cytochrome P450(CYP)gene family in plant tolerance to THMMs has been recently highlighted,but there is still a lack of comprehensive understanding,especially in relation to metabolites.This study delved into the identification of CYP genes that are linked to the tolerance mechanisms of plants in response to heavy metal stress.The findings highlight the significant metabolic pathways that contribute to this resilience,using rice and Arabidopsis as exemplars.THMM exposure changed CYP gene expression in plants,and THMM antidotes mitigated its downregulation and that of flavonoid biosynthesis genes.CYP genes involved in THMM responses were predominantly enriched in the pathways associated with flavonoid synthesis,indicating functional adaptations to distinct stresses.Notably,anthocyanin(Ant)accumulation,a type of flavonoid,affected the uptake of various heavy metals in Brassica rapa,with flavonoid biosynthesis-associated genes correlating with Cd or As tolerance and Ant content.These findings highlight the critical importance of flavonoid metabolism and the intricate network of biosynthesis genes in bolstering plant resilience against heavy metal stress.This enhanced understanding paves the way for significant advancements in phytoremediation technologies,offering innovative solutions for soil and water decontamination.
文摘细胞色素P450酶是自然界最大的一类氧化还原酶超家族,广泛参与药物代谢、植物次生代谢以及环境污染物转化等重要生物过程。为了生成具有高保真度和多样性的人工P450酶序列,本研究提出了一种基于扩散模型的P450酶序列生成方法——P450Diff2。该模型基于参数量达6.4亿的EvoDiff-Seq框架进行构建,并以来自NCBI、GMind注释、RNA-Seq组装和宏基因组数据库的1041254条非冗余P450蛋白序列作为训练数据。基于训练完成的模型进行序列生成评估,结果显示,P450Diff2在氨基酸组成分布、序列特征空间、序列相似性分布以及结构合理性等多个维度均优于此前提出的P450Diffusion模型,其所生成序列的平均局部预测置信度(predicted local distance difference test,pLDDT)达到72.29。实验结果进一步表明,所生成的序列中有60%能够正确折叠为具有生物活性的P450酶,表明本方法不仅有效保留了天然序列的结构特征,同时具备一定的功能性序列生成可能性。结合大规模序列生成与筛选流程,有望快速设计生成高效新酶序列,降低实验筛选的时间与成本,为新酶的高效、可扩展设计提供了全新的参考。
