In our previous EPSRC-NSF funded project, we developed an innovative, versatile integrase enzyme based system for assembly and rearrangement of DNA modules – the “Syntegron technology platform” that allows rapid and efficient assembly of multiple standardized DNA modules into large assemblies such as metabolic pathways and genetic circuits, and to exchange individual parts of assemblies (e.g., regulatory elements, homologous or mutant genes) to allow variation and optimization. Here, we propose to build on these core technologies to develop a comprehensive experimental and computational toolkit for the synthetic biology community, enabling the rapid generation of biosynthetic industrially and medically-relevant products. We will pursue and demonstrate these capabilities through two complementary and interrelated program objectives: Objective 1: Develop Syntegron “parts” enabling the discovery and production of diverse, valuable natural products. An important source for drug leads has been plant natural products – up to 60% of the successful drugs are of natural origin but many of these drugs are produced in miniscule amounts in their native hosts, making the drugs expensive and environmentally taxing to harvest. Organic chemistry methodologies have been widely used to synthesize pharmaceuticals in current use but many natural/nature-inspired pharmaceutically-relevant scaffolds cannot be achieved by these methods. An alternative is the use of enzymes for combinatorial synthesis inside the cell, to enable the production of drug candidates from inexpensive, green starting materials. The goal of this research is to synthesize a variety of natural product variants e.g. terpenes using the Syntegron technology platform constructed in the first phase of funding, and to manufacture these natural product variants for drug discovery applications. Specifically, we will 1) Take advantage of the recently described phenomena of plant metabolic gene clusters to identify genes encoding enzymes involved in plant secondary metabolite producing pathways via the development of novel bioinformatic and data mining tools. 2) We will then use the Syntegron platform to assemble the synthetic metabolic pathways for natural products e.g. the triterpenes and enzymes that will decorate the natural products with various functional groups to synthesize a variety of natural product variants. 3) Develop new Syntegron host strains for metabolic engineering 4) Engineer in vivo biosensors for natural products and their variants and use these sensors in high throughput screens allowing dynamic optimization of metabolic pathways.
|Effective start/end date||9/1/13 → 8/31/16|
- National Science Foundation (MCB-1341414)
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