Producing the Polyketide Antibiotic Actinorhodin From Azo Dye Waste: In silico Design and Optimization of a Synthetic Pathway in Pseudomonas putida KT2440
ISEF · 2022 Biomedical Engineering
Overview
Recent advances in synthetic biology have enabled the conversion of industrial wastes into high-value chemical products, including specialized metabolites such as polyketides. Azo dyes, a major pollutant commonly discharged by food, textile, and pharmaceutical industries, have the potential to be repurposed as a feedstock for biomanufacturing. Here, I sought to design a synthetic pathway in Pseudomonas putida KT2440 capable of recycling azo compounds into the polyketide actinorhodin (ACT). The synthetic pathway was constructed by combining a set of reactions identified from publicly-available databases. Flux balance analysis was used to test the metabolic potential and limitations of the synthetic pathway in P. putida using a modified version of the iJN1463 genome-scale metabolic model. The algorithms Optknock, Optgene, and FSEOF were employed to examine whether coupling dye consumption and ACT production with growth could be a suitable optimization strategy. It was revealed that while dye degradation is always coupled with growth, growth-coupling for ACT production is theoretically inefficient. Thus, a synthetic genetic circuit was constructed to decouple growth and ACT production in a two-stage fermentation scenario. Five refactored operons were designed to modularize the ACT gene cluster and avoid overlapping of native and synthetic regulatory elements. To express the system in P. putida, the regulatory circuit and refactored operons were integrated into pRO1600 Pseudomonas-specific and pBBR1 broad-host-range plasmids using Gibson Assembly. This research demonstrates that the conversion of azo dyes into actinorhodin in P. putida is feasible. Moreover, this work establishes a systematic workflow and testable genetic designs to be further investigated experimentally.
Competition history
- ISEF 2022
Resources
Related projects
ISEF · 2026
Designing Safer Microbial Plastic Degraders: CasTune — An Optimized dCas13d-IF3 Framework for Higher PETase Production in E. coli
ISEF · 2015
Azoreductase Activity: Bioremediation for Treatment of Synthetic Diazo Dyes in Polluted Water
ISEF · 2014
The Conjugative Plasmid RK2 as a Delivery System for Artificial AnatheriaH Genes: A Novel Synthetic Biology Alternative to Traditional Antibiotics
ISEF · 2017
Improving Nitration Activity of Fused TxtE-CYP102AI Reductase Domain by Optimizing the Linker Length
Closest projects by meaning, across every fair and year in the corpus.
Source: Regeneron International Science and Engineering Fair