Microbial Strigolactone Biosynthesis via E. coli-Yeast Co-Culture
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Solution Overview
Problem
The extremely low abundance of strigolactones in nature, challenging chemical synthesis, and incomplete structure-function correlation hinder the development of commercial agricultural applications for strigolactones.
Innovation Solution
A bacterial and yeast co-culture system is developed to de novo biosynthesize non-canonical and canonical strigolactones by splitting the biosynthetic pathway into two modules: CL production in E. coli and SL synthesis in yeast, using engineered strains expressing specific enzymes such as DWARF27, CCD7, CCD8, cytochrome P450 reductase, and synthetase genes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If chemical synthesis is used to produce strigolactones, then synthesized SL standards and analogues can be provided, but the synthetic analogs are generally less active than natural SLs and the process is laborious and expensive
Solution Approach 1:
The biosynthetic pathway is segmented into two distinct modules: Module 1 in E. coli for carlactone production and Module 2 in yeast for strigolactone synthesis. This segmentation allows each organism to be optimized for its specific function, simplifying the overall manufacturing process while maintaining high productivity and natural SL activity.
Solution Approach 2:
Carlactone serves as an intermediary compound that is produced by E. coli and then consumed by yeast to synthesize strigolactones. This intermediary approach enables the division of labor between two organisms, allowing each to specialize in a specific biosynthetic step and improving overall efficiency.
2Adaptability or versatility
If de novo synthesis of canonical SLs in a microbial host is attempted, then a microbial platform for SL production can be established, but the complete biosynthetic pathway has not been successfully reconstructed
Solution Approach 1:
The complete SL biosynthetic pathway is segmented and distributed across two different microbial hosts: E. coli for the carlactone production module and yeast for the strigolactone synthesis module. This segmentation enables successful de novo synthesis by allowing each organism to express the specific enzyme set needed for its designated function, overcoming the limitation of attempting to reconstruct the entire pathway in a single host.
Solution Approach 2:
The invention creates a universal microbial production platform by combining E. coli and yeast systems that can collectively perform the complete SL biosynthetic pathway. This multi-functional approach allows the system to produce both non-canonical and canonical SLs, providing versatility while maintaining reliability through specialized enzymatic functions in each organism.
3Adaptability or versatility
If the biosynthetic pathway is split into two modules for co-culture production, then both non-canonical and canonical SLs can be synthesized, but the system complexity increases
Solution Approach 1:
The biosynthetic pathway is segmented into two modules expressed in different organisms (E. coli and yeast), which simplifies the enzymatic requirements for each host while enabling comprehensive SL production. This segmentation reduces individual organism complexity while achieving high production versatility.
Solution Approach 2:
Carlactone acts as an intermediary that facilitates the transfer of metabolic flux between the two organisms. E. coli produces carlactone which is then consumed by yeast for strigolactone synthesis, creating a streamlined co-culture system that manages complexity through a clear functional division and intermediate compound exchange.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enables the stable production of strigolactones, including carlactone, carlactonic acid, 5-deoxystrigol, 4-deoxyorobanchol, and orobanchol, with titers up to 15 μg/L, providing a viable microbial platform for agricultural applications.
Implementation Method 1
de novo synthesis of CL was achieved in the β-carotene-accumulating strains of E. coli by introducing an isomerase DWARF27 (D27) and two carotenoid cleavage dioxygenases (CCD7 and CCD8, respectively)
Implementation Method 2
The biosynthetic pathway from CL to SLs (including 5DS, 4DO and orobanchol) was achieved by expressing various cytochrome P450s and the corresponding reductase in yeast strain
Implementation Method 3
expressing various cytochrome P450s and the corresponding reductase in yeast strain
Implementation Method 4
a bacterial and yeast, e.g., E. coli-S. cerevisiae, co-culture strategy to generate a microbial SL platform for synthesizing both non-canonical and canonical SLs
Data Source
AI summary
The present disclosure provides a bacterial and yeast co-culture system and methods of making and using such co-culture systems for producing strigolactones.