Toward engineering synthetic microbial metabolism

Abstract

The generation of well-characterized parts and the formulation of biological design principles in synthetic biology are laying the foundation for more complex and advanced microbial metabolic engineering. Improvements in de novo DNA synthesis and codon-optimization alone are already contributing to the manufacturing of pathway enzymes with improved or novel function. Further development of analytical and computer-aided design tools should accelerate the forward engineering of precisely regulated synthetic pathways by providing a standard framework for the predictable design of biological systems from well-characterized parts. In this review we discuss the current state of synthetic biology within a four-stage framework (design, modeling, synthesis, analysis) and highlight areas requiring further advancement to facilitate true engineering of synthetic microbial metabolism.

Figure 1

Synthetic biology four-step engineering workflow. The design and modeling steps (white text) are computational in nature, whereas the synthesis and analysis steps (black text) are experimentally-based. The workflow presents the relationships between the individual steps as well as a logical progression using a synthetic biology framework to generate a product that meets determined specifications.

Figure 2

Cartoon of an engineered cell. Four distinct strategies have been combined to illustrate a synthetic biology approach to metabolic engineering. The engineered pathway is encoded by a synthetic operon designed from standard parts/components including tunable intergenic regions (TIGRs) that provide differential control over mRNA stability [41]. An upstream device/module, a transcriptional modular AND gate acts as a master regulator by requiring two separate inputs to turn on expression of the pathway [15]. The enzymes have been tagged with a peptide ligand that binds them to a synthetic protein scaffold, forming a complex of colocalized pathway enzymes [27]. The central metabolism of the host/chassis has been modified to interface with the engineered system/network, providing a sufficient flux of necessary precursors and cofactors as well as increased transporters to facilitate product secretion [7]. This image was generated in TinkerCell.

Figure 3

Convergence of the different realms of synthetic biology. In addition to efforts to refine and characterize genetic parts based on DNA, research on expanding the genetic code and implementing biological processes in cell-free systems will likely integrate with metabolic engineering projects in the future. Genome engineering and synthetic genomics will enable the manufacture of customizable minimal genomes. Alternatively, protocell engineering may provide a non-biological chassis to house synthetic chemistry.