Microbial Synthetic Epigenetic Tools Design and Applications.

Microbial synthetic epigenetics offers significant opportunities for the design of synthetic biology tools by leveraging reversible gene control mechanisms without altering DNA sequences. However, limited understanding and a lack of technologies for thorough analysis of the mechanisms behind epigenetic modifications have hampered their utilization in biotechnological applications. In this review, we explore advancements in developing epigenetic-based synthetic gene regulatory tools at both transcriptional and post-transcriptional levels. Furthermore, we examine strategies developed to construct epigenetic-based circuits that provide controllable and stable gene regulation, aiming to boost the performance of microbial chassis cells. Finally, we discuss the current challenges and perspectives in the development of synthetic epigenetic tools.

Synthetic epigenetics-assisted microbial chassis engineering.

Microbial chassis engineering is the milestone of efficient biotechnological applications. However, microbial chassis cell engineering is adversely affected by (i) regulatory tool orthogonality, (ii) host metabolic fitness, and (iii) cell population heterogeneity. Herein, we explore how synthetic epigenetics can potentially address these limitations and offer insights into prospects in this field.

Bifunctional optogenetic switch for improving shikimic acid production in E. coli.

BACKGROUND: Biomass formation and product synthesis decoupling have been proven to be promising to increase the titer of desired value add products. Optogenetics provides a potential strategy to develop light-induced circuits that conditionally control metabolic flux redistribution for enhanced microbial production. However, the limited number of light-sensitive proteins available to date hinders the progress of light-controlled tools. RESULTS: To address these issues, two optogenetic systems (TPRS and TPAS) were constructed by reprogramming the widely used repressor TetR and protease TEVp to expand the current optogenetic toolkit. By merging the two systems, a bifunctional optogenetic switch was constructed to enable orthogonally regulated gene transcription and protein accumulation. Application of this bifunctional switch to decouple biomass formation and shikimic acid biosynthesis allowed 35 g/L of shikimic acid production in a minimal medium from glucose, representing the highest titer reported to date by E. coli without the addition of any chemical inducers and expensive aromatic amino acids. This titer was further boosted to 76 g/L when using rich medium fermentation. CONCLUSION: The cost effective and light-controlled switch reported here provides important insights into environmentally friendly tools for metabolic pathway regulation and should be applicable to the production of other value-add chemicals.