Abstract
Heterologous expression of biosynthetic gene clusters (BGCs) avails yield improvements and mining of natural products, but it is limited by lacking of more efficient Gram-negative chassis. The proteobacterium Schlegelella brevitalea DSM 7029 exhibits potential for heterologous BGC expression, but its cells undergo early autolysis, hindering further applications. Herein, we rationally construct DC and DT series genome-reduced S. brevitalea mutants by sequential deletions of endogenous BGCs and the nonessential genomic regions, respectively. The DC5 to DC7 mutants affect growth, while the DT series mutants show improved growth characteristics with alleviated cell autolysis. The yield improvements of six proteobacterial natural products and successful identification of chitinimides from Chitinimonas koreensis via heterologous expression in DT mutants demonstrate their superiority to wild-type DSM 7029 and two commonly used Gram-negative chassis Escherichia coli and Pseudomonas putida. Our study expands the panel of Gram-negative chassis and facilitates the discovery of natural products by heterologous expression.
Highlights
Heterologous expression of biosynthetic gene clusters (BGCs) avails yield improvements and mining of natural products, but it is limited by lacking of more efficient Gram-negative chassis
During our deep investigation of this strain to improve the production of epothilones, we found that the early autolysis of DSM 7029 led to most cell death after 48 h of fermentation, which has been recently reported to occur in many Gram-negative bacteria[38,39,40], severely affecting the biomass of DSM 7029 and restricting the heterologous production yields of desired compounds[41]
We recently established an electroporation procedure and efficient genetic engineering system based on the bacteriophage recombinase pair in this strain, dramatically facilitating precise insertion of potent promoters to drive cryptic BGCs for discovery of natural products produced by strain DSM 7029 and those produced by other Burkholderiales species and providing a fluent technological stage for further optimization[46,47,48]
Summary
Heterologous expression of biosynthetic gene clusters (BGCs) avails yield improvements and mining of natural products, but it is limited by lacking of more efficient Gram-negative chassis. The β-proteobacterial strain Schlegelella brevitalea DSM 7029, formerly known as Polyangium brachysporum K481-B101 or Burkholderiales strain DSM 7029, was previously utilized as a heterologous host to produce proteobacterial nonribosomal peptide/polyketide (NRP/PK) natural products with relatively high yields, such as epothilone and vioprolide from myxobacteria (δ-proteobacteria) and rhizomide from Burkholderiales (βproteobacteria)[33,34,35] This strain possesses essential biosynthetic elements, e.g., 4′-phosphopantetheinyl transferase, and generates the important PK extender unit methylmalonyl-CoA that is not produced at a detectable level in another commonly used Gramnegative chassis, Pseudomonas putida[33,36], for the efficient biosynthesis of PKs and NRPs37. We discovered and characterized several strong constitutive promoters in strain DSM 7029, which are beneficial for the optimization of the yield of important metabolites in this chassis[34]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
