A new E. coli-based system uses metabolic engineering and co-cultures to produce various halogenated tryptophan molecules from glucose, providing an environmentally friendly alternative for creating diverse compounds used in pharmaceuticals and other industries.
Halogenated molecules are critical to various industries, including pharmaceuticals, agrochemicals, and materials science, due to their enhanced biological activities and stability. The precise incorporation of halogen atoms into organic compounds can significantly improve their efficacy, selectivity, and metabolic stability, making them valuable for the development of advanced drugs and specialized materials.
As the demand for such tailored compounds grows, there is an increasing need for sustainable and efficient methods to produce a diverse array of halogenated derivatives. Biological synthesis, leveraging enzymatic processes, offers a promising alternative to traditional chemical methods by providing greater specificity while reducing environmental impact. However, current approaches to synthesizing halogenated molecules face significant challenges.
Traditional chemical synthesis often relies on toxic reagents and harsh conditions, leading to poor atom economy and difficulties in achieving regio- and stereo-selectivity. These methods can also generate substantial waste, making them less environmentally friendly. Existing biosynthetic techniques may also suffer from limited enzyme efficiency, narrow substrate scope, and restricted halogenation sites, constraining the diversity and yield of the desired halogenated products. Additionally, metabolic bottlenecks in host organisms can impede the scalable production of these valuable compounds, highlighting the need for more advanced and versatile biosynthetic platforms.
UT researchers developed a technology for the de novo biosynthesis of various halogenated tryptophan-derived molecules in E. coli, utilizing glucose as the starting material. This system integrates metabolic engineering with a modular co-culture approach to optimize production efficiency and molecular diversity. Key features include the deletion of tnaA and trpR genes to prevent tryptophan degradation and repression, the introduction of feedback-resistant mutations in critical biosynthetic genes, and the overexpression of enzymes involved in tryptophan synthesis.
Additionally, the platform employs specific halogenases alongside a thermostable flavin reductase to achieve precise halogenation of tryptophan at multiple positions. By segregating the production and conversion processes into distinct E. coli strains, the system successfully synthesizes 26 distinct halogenated molecules, including novel beta-carbolines and prodrug precursors, offering a sustainable alternative to traditional chemical methods.
This technology stands out due to its innovative combination of metabolic engineering and modular co-culture strategies, enabling the efficient production of a wide range of halogenated compounds that are challenging to synthesize chemically. The precise regioselectivity of halogenase enzymes ensures high specificity in halogenation, while the use of feedback-resistant mutations and overexpressed biosynthetic pathways maximizes precursor availability. The thermostable flavin reductase enhances the efficiency of the halogenation process, and the spatial separation of production and conversion in different strains minimizes metabolic burden and increases overall yield. This approach not only enhances the diversity and scalability of bio-based halogenated molecule production but also promotes environmentally friendly practices, making it highly valuable for pharmaceuticals, agrochemicals, and materials science applications.
U.S. Provisional filed 12 Sept 2024
https://www.nature.com/articles/s41467-023-40242-9