Abstract
BackgroundIndigo is a color molecule with a long history of being used as a textile dye. The conventional production methods are facing increasing economy, sustainability and environmental challenges. Therefore, developing a green synthesis method converting renewable feedstocks to indigo using engineered microbes is of great research and application interest. However, the efficiency of the indigo microbial biosynthesis is still low and needs to be improved by proper metabolic engineering strategies.ResultsIn the present study, we adopted several metabolic engineering strategies to establish an efficient microbial biosynthesis system for converting renewable carbon substrates to indigo. First, a microbial co-culture was developed using two individually engineered E. coli strains to accommodate the indigo biosynthesis pathway, and the balancing of the overall pathway was achieved by manipulating the ratio of co-culture strains harboring different pathway modules. Through carbon source optimization and application of biosensor-assisted cell selection circuit, the indigo production was improved significantly. In addition, the global transcription machinery engineering (gTME) approach was utilized to establish a high-performance co-culture variant to further enhance the indigo production. Through the step-wise modification of the established system, the indigo bioproduction reached 104.3 mg/L, which was 11.4-fold higher than the parental indigo producing strain.ConclusionThis work combines modular co-culture engineering, biosensing, and gTME for addressing the challenges of the indigo biosynthesis, which has not been explored before. The findings of this study confirm the effectiveness of the developed approach and offer a new perspective for efficient indigo bioproduction. More broadly, this innovative approach has the potential for wider application in future studies of other valuable biochemicals’ biosynthesis.
Highlights
Indigo is an aromatic molecule with deep blue color and has been historically used as a natural dye
Developing an E. coli–E. coli co‐culture for the indigo biosynthesis on glucose and glycerol The indigo biosynthetic pathway consists of tryptophan provision and its conversion to indigo (Fig. 1)
The best co-culture variant, BTR1: BRM-353, produced 80.6 mg/L indigo from 5 g/L glycerol after 48 h cultivation, which was 135% higher than the BTR1:BRC control co-culture. These findings suggested that desirable phenotypes for high indigo biosynthesis in the co-culture were successfully created using global transcription machinery engineering (gTME)
Summary
Indigo is an aromatic molecule with deep blue color and has been historically used as a natural dye. Most of the indigo used in industry is currently obtained by either plant extraction or chemical synthetic process [1]. These methods need to use massive amount of chemicals as extractant, reactant or catalyst, leading to a series of process economic, sustainability, and environmental problems. To address this challenge, the indigo biosynthesis using renewable materials has been explored as a robust alternative. The efficiency of the indigo microbial biosynthesis is still low and needs to be improved by proper metabolic engineering strategies
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