Fig. 2
From: Cell-free prediction of protein expression costs for growing cells
Predictive model for resource competition between a synthetic construct and capacity monitor. a Schematic of mathematical model of competitive translation between capacity monitor construct and a construct expressing a GoI, with a finite ribosome pool. A free ribosome binds to an unoccupied RBS at a rate a+ (a+ = a+M × RBS strength) and either unbinds and returns to the free ribosome pool at a rate a− (a− = a−M/RBS strength), or initiates synthesis at a rate b0 (b0 = b0M × RBS strength). Once synthesis has initiated, all the mechanisms of the protein synthesis are gathered in the lumped parameter γ, which represents the cost of protein synthesis. The number of protein synthesis steps depends on size (mRNA size/30—as 30 bases represents the footprint of a ribosome). Each synthesis step is considered to proceed with the same cost γ. b Heat maps of simulated capacity monitor expression (monitor output) when mRNA size and γ value of a synthetic construct are varied. The heat map is used to determine the γ value of each construct in Fig. 1f. c A new heat map simulated for weaker RBS is used to predict monitor output using γ values calculated in b. As prediction is done for cell lysate, in vivo predictions are then deduced from the relationship between cell lysate and in vivo measurements as per Fig. 1f. d First row: heat maps of simulated capacity (monitor output) when mRNA concentration and γ-values of a synthetic circuit are varied. Each heat map is a construct with different RBS strength. Second row: capacity (monitor output) as a function of mRNA level. Each line corresponds to different γ-values. e Measurements of normalised in vitro capacity when increasing synthetic construct DNA is added to cell lysate. Top: mkate with a strong RBS. Bottom: viob-mkate with a strong RBS. The calculated γ and RBS strengths (from Supplementary Fig. 4B) are shown for each construct. Lines show a fit to data points. Error bars show standard error of three independent repeats. Values are normalised to the capacity obtained with capacity monitor plasmid alone
