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A combined strategy for the overproduction of complex ergot alkaloid agroclavine

Synthetic and Systems Biotechnology 2022年第4期7卷 1126-1132页

作者： Zhi-Pu Yu Chunyan An Yongpeng Yao Chang-Yun Wang Zhoutong Sun Chengsen Cui Ling Liu Shu-Shan Gao Key Laboratory of Marine Drugs The Ministry of Education of ChinaInstitute of Evolution&Marine BiodiversitySchool of Medicine and PharmacyOcean University of ChinaQingdao266003PR China Tianjin Institute of Industrial Biotechnology Chinese Academy of SciencesTianjin300308PR China Laboratory for Marine Drugs and Bioproducts Qingdao National Laboratory for Marine Science and TechnologyQingdao266003PR China State Key Laboratory of Mycology Institute of MicrobiologyChinese Academy of SciencesBeijing100101PR China Beijing Institute for Drug Control NMPA Key Laboratory for Research and Evaluation of Generic DrugsBeijing Key Laboratory of Analysis and Evaluation on Chinese MedicineBeijing102206PR China National Technology Innovation Center of Synthetic Biology Tianjin300308PR China

Microbial cell factories(MCFs)and cell-free systems(CFSs)are generally considered as two unrelated approaches for the biosynthesis of biomolecules.In the current study,two systems were combined together for the overproduction of agroclavine(AC),a structurally complex ergot alkaloid.The whole biosynthetic pathway for AC was split into the early pathway and the late pathway at the point of the FAD-linked oxidoreductase EasE,which was reconstituted in an MCF(Aspergillus nidulans)and a four-enzyme CFS,respectively.The final titer of AC of this combined system is 1209 mg/L,which is the highest one that has been reported so far,to the best of our knowledge.The development of such a combined route could potentially avoid the limitations of both MCF and CFS systems,and boost the production of complex ergot alkaloids with polycyclic ring systems.

关键词： Microbial cell factories Cell-free systems Overproduction Agroclavine ergot alkaloids

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Efficient genome editing in Claviceps purpurea using a CRISPR/Cas9 ribonucleoprotein method

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Synthetic and Systems Biotechnology 2022年第2期7卷 664-670页

作者： Lu Yu Meili Xiao Zhihua Zhu Yinmei Wang Zhihua Zhou Pingping Wang Gen Zou CAS-Key Laboratory of Synthetic Biology CAS Center for Excellence in Molecular Plant SciencesInstitute of Plant Physiology and EcologyChinese Academy of Sciences300 Fenglin RdShanghai200032China University of Chinese Academy of Sciences Beijing100049China Shanghai Key Laboratory of Agricultural Genetics and Breeding Institute of Edible FungiShanghai Academy of Agriculture Science1000 Jinqi RdFengxianShanghai201403China

Claviceps purpurea produces many pharmacologically important ergot alkaloids(EAS),which are widely used to treat migraine and hypertension and to aid childbirth.Although an EAS biosynthetic cluster of C.purpurea has been discovered more than 20 years ago,the complete biosynthetic pathway of EAS has not been fully characterized until now.The main obstacle to elucidating this pathway and strain modification is the lack of efficient genome-editing tools for C.purpurea.The conventional gene manipulation method for C.purpurea relies on homologous recombination(HR),although the efficiency of HR in C.purpurea is very low(~1-5%).Consequently,the disruption of target genes is laborious and time-consuming.Although CRISPR/Cas9 genome-editing methods based on in vivo Cas9 expression and gRNA transcription have been reported recently,their gene-disruption efficiency is still very low.Here,we developed an efficient genome-editing system in C.purpurea based on in vitro assembled CRISPR/Cas9 gRNA ribonucleoprotein complexes.As proof of principle,three target genes were efficiently knocked out using this CRISPR/Cas9 ribonucleoprotein complex-mediated HR system,with editing efficiencies ranging from 50%to 100%.Inactivation of the three genes,which are closely related to uridine biosynthesis(ura5),hypha morphology(rac),and EAS production(easA),resulted in a uridine auxotrophic mutant,a mutant with a drastically different phenotype in axenic culture,and a mutant that did not produce EAS,respectively.Our ribonucleoprotein-based genome-editing system has a great advantage over conventional and in vivo CRISPR/Cas9 methods for genome editing in C.purpurea,which will greatly facilitate elucidation of the EAS biosynthetic pathway and other future basic and applied research on C.purpurea.

关键词： Ribonucleoprotein Genome editing ergot alkaloids Biosynthetic pathway Homologous recombination

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Total and formal synthesis of some ergot alkaloids

Total and formal synthesis of some ergot alkaloids

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中国化学会第十二届全国天然有机化学学术会议

作者： Wei Wang Yang Mi Jin-Quan Chen Jia-Tian Lu Zi-Fa Shi Xiao-Ping Cao State Key Laboratory of Applied Organic Chemistry College of Chemistry and Chemical Engineering Lanzhou University

The ergot family represents a significant class of indole alkaloids with broad pharmacological activity.Most of the ergot alkaloids are isolated from various Claviceps species,although they can also be isolated from o... 详细信息

关键词： ergot alkaloids Total and formal synthesis

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