Enhanced Heterologous Spinosad Production from a 79-kb Synthetic Multioperon Assembly

Research output: Contribution to journalResearch articleContributedpeer-review

Contributors

  • Chaoyi Song - (Author)
  • Ji Luan - (Author)
  • Qingwen Cui - (Author)
  • Qiuyue Duan - (Author)
  • Zhen Li - (Author)
  • Yunsheng Gao - (Author)
  • Ruijuan Li - (Author)
  • Aiying Li - (Author)
  • Yuemao Shen - (Author)
  • Yuezhong Li - (Author)
  • A. Francis Stewart - , Chair of Applied Genomics, GenArc GmbH, Dresden (Author)
  • Youming Zhang - (Author)
  • Jun Fu - (Author)
  • Hailong Wang - (Author)

Abstract

Refactoring biosynthetic pathways for enhanced secondary metabolite production is a central challenge for synthetic biology. Here we applied advanced DNA assembly methods and a uniform overexpression logic using constitutive promoters to achieve efficient heterologous production of the complex insecticidal macrolide spinosad. We constructed a 79-kb artificial gene cluster in which 23 biosynthetic genes were grouped into 7 operons, each with a strong constitutive promoter. Compared with the original gene cluster, the artificial gene cluster resulted in a 328-fold enhanced spinosad production in Streptomyces albus J1074. To achieve this goal, we applied the ExoCET DNA assembly method to build a plasmid from 13 GC-rich fragments with high efficiency in one step. Together with our previous direct cloning and recombineering tools, we present new synthetic biology options for refactoring large gene clusters for diverse applications.

Details

Original languageEnglish
Pages (from-to)137-147
Number of pages11
JournalACS synthetic biology
Volume8
Issue number1
Publication statusPublished - 18 Jan 2019
Peer-reviewedYes

External IDs

PubMed 30590919
ORCID /0000-0002-4754-1707/work/142248102

Keywords

Keywords

  • biosynthesis, DNA engineering, gene clusters, heterologous expression, secondary metabolites, synthetic operons