Autonomous translation modulation in a homeostatic artificial cell
Biophysics
Statistical Physics, Nonlinear Dynamics, Complex Systems, Soft and Fluid Matter, Biological Physics
Final Report Abstract
All the molecules that make up a cell, such as a bacterium, are self-regenerated. This separates them from nutrients. Self-regeneration is the remaking of a lost part with an analogous and functional copy. It is a fundamental property of organisms as it forms the basis for self-replication and homeostasis. This self-maintaining capability is still missing from engineered biomimetic systems. The field of bottom-up synthetic biology is constructing biomimetic systems from molecules such as DNA, RNA, and proteins to re-create features from biological systems outside of cells. For example, it is possible to reconstitute gene expression from DNA templates outside cells using only purified components. The list of components required to achieve gene expression contains 36 proteins, ribosomes, amino acids, and tRNA and is called the PURE (protein synthesis using recombinant elements) system. Before self-regeneration of the PURE is achievable, the synthesis and functionality of all the components must be verified. We identified 30 of the 36 proteins directly involved in translation and sought to prove their synthesis and functionality in the PURE system. We studied the expression of PURE proteins in a model synthetic cell that is realized in micrometre sized compartments on a silicon chip with DNA molecules attached to the surface. We established a recovery assay that works in a delta PURE system devoid of one of the 30 proteins of interest. By adding the DNA coding for that protein, we monitor the recovery of gene expression capabilities due to the freshly in vitro synthesized protein. We were thus able to show that the proteins involved in translation can be synthesized in vitro and are functional, laying the foundations for a self-regenerating cell-free gene expression system within a synthetic cell.
