Abstract
BackgroundSynthetic biology is used to develop cell factories for production of chemicals by constructively importing heterologous pathways into industrial microorganisms. In this work we present a retrosynthetic approach to the production of therapeutics with the goal of developing an in situ drug delivery device in host cells. Retrosynthesis, a concept originally proposed for synthetic chemistry, iteratively applies reversed chemical transformations (reversed enzyme-catalyzed reactions in the metabolic space) starting from a target product to reach precursors that are endogenous to the chassis. So far, a wider adoption of retrosynthesis into the manufacturing pipeline has been hindered by the complexity of enumerating all feasible biosynthetic pathways for a given compound.ResultsIn our method, we efficiently address the complexity problem by coding substrates, products and reactions into molecular signatures. Metabolic maps are represented using hypergraphs and the complexity is controlled by varying the specificity of the molecular signature. Furthermore, our method enables candidate pathways to be ranked to determine which ones are best to engineer. The proposed ranking function can integrate data from different sources such as host compatibility for inserted genes, the estimation of steady-state fluxes from the genome-wide reconstruction of the organism's metabolism, or the estimation of metabolite toxicity from experimental assays. We use several machine-learning tools in order to estimate enzyme activity and reaction efficiency at each step of the identified pathways. Examples of production in bacteria and yeast for two antibiotics and for one antitumor agent, as well as for several essential metabolites are outlined.ConclusionsWe present here a unified framework that integrates diverse techniques involved in the design of heterologous biosynthetic pathways through a retrosynthetic approach in the reaction signature space. Our engineering methodology enables the flexible design of industrial microorganisms for the efficient on-demand production of chemical compounds with therapeutic applications.
Highlights
Synthetic biology is used to develop cell factories for production of chemicals by constructively importing heterologous pathways into industrial microorganisms
Synthetic biology is being used for therapeutic production either to develop cell factories using industrial microorganisms [1,2] or to synthesize genetic circuits allowing in situ therapeutic delivery [3]
We present here a unified framework that combines several techniques involved in the design of heterologous biosynthetic pathways through a retrosynthetic approach in the reaction signature space, enabling the flexible design of industrial microorganisms for the efficient on-demand production of chemical compounds of interest
Summary
Synthetic biology is used to develop cell factories for production of chemicals by constructively importing heterologous pathways into industrial microorganisms. Synthetic biology is being used for therapeutic production either to develop cell factories using industrial microorganisms [1,2] or to synthesize genetic circuits allowing in situ therapeutic delivery [3]. Recombinant DNA technology has already provided the ability to genetically engineer cell strains in order to import pathways from other organisms capable of producing small molecule chemicals into microbial chassis. Many successful examples of bioproduction of chemicals with therapeutic interest through metabolic engineering have been reported. Heterologous production of therapeutically important antibiotics such as aminoglycosides derivatives, which include ribostamycin [9], neomycin, gentamicin and kanamycin, as well as other natural products like polyketides [10,11] and nonribosomal peptides [12] have been reported. Flexible production of novel antibiotics is of special interest in order to fight against the increasing emergence of multidrug-resistant pathogens [13,14,15]
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