Synthetic Biology Outside the Cell: Linking Computational Tools to Cell-Free Systems

Abstract

As mathematical models become more commonly integrated into the study of biology, a common language for describing biological processes is manifesting. Many tools have emerged for the simulation of in vivo synthetic biological systems, with only a few examples of prominent work done on predicting the dynamics of cell-free synthetic systems. At the same time, experimental biologists have begun to study dynamics of in vitro systems encapsulated by amphiphilic molecules, opening the door for the development of a new generation of biomimetic systems. In this review, we explore both in vivo and in vitro models of biochemical networks with a special focus on tools that could be applied to the construction of cell-free expression systems. We believe that quantitative studies of complex cellular mechanisms and pathways in synthetic systems can yield important insights into what makes cells different from conventional chemical systems.

Metabolic Perceptrons for Neural Computing in Biological Systems

“…Finally, 6 using our integrated model trained on the cell-free metabolic circuits we build a more 7 sophisticated device called the "metabolic perceptron", which allows desired binary 8 classification of multi-input metabolite combinations by applying model-predicted weights 9 on the input metabolites before analog addition, and demonstrate its utility through two 10 examples of four-input binary classifiers. Altogether, in this work we demonstrate the 11 potential of synthetic metabolic circuits, along with model-assisted design, to perform 12 complex computations in biological systems. 13…”

“…Increasing the concentration 9 of the enzymes produces a higher amount of benzoate from the input metabolites, and 10 hence higher GFP fluorescence. Compared to the others, the hippurate transducer 11 reached higher GFP expression at a given concentration of the enzyme and the input, 12 and biphenyl-2,3-diol reached the weakest signal. For the biphenyl-2,3-diol transducer 13 built with two enzymes (Figure 4e), both enzymes are added at the same concentration 14 (e.g., 1 nM of "enzyme DNA" indicates 1 nM each of plasmids encoding enzymes bphC 15 and bphD).…”

“…This model (presented in gray lines in Figure 1d,e,f and Figure 2c), 10 which was not trained on adder data but only on actuator, transducer, and transducers 11 with resource competition data, recapitulates it well. This indicates that the model 12 accounts for all important effects underlying the data. The full training process is 13 presented in the Methods section, and a table summarising scores of estimated goodness 14 of fit of our model is presented in Supplementary Table S1.…”

“…A Whole-cell metabolic concentration adder 10 A metabolic concentration adder is a device composed of more than one transducer that 11 converts their respective input metabolites into a common transduced output metabolite. 12 For our whole-cell concentration adder, we combined two transducers to build a 13 hippurate-benzaldehyde adder actuated by the benzoate circuit (Figure 2a). Unlike digital 14 bit-adders that exhibit an ON-OFF digital behavior, our metabolic adders exhibit a 15 continuous analog behavior that is natural for metabolic signal conversion 51 (Figure 2b 16 and Supplementary Figure S3) .…”

“…Finally, cell-free systems are highly adjustable for different 11 performance parameters and components. In all, these advantages of cell-free systems 12 enable us to develop more complex computations than the whole-cell adder. 13 14 Following from our recent work 56 , we first characterized a cell-free benzoate actuator to 15 be used downstream of other metabolic transducers.…”