Fig. 3: Feedforward ribosome controller compensates for burden caused by a GOI (RFP) activation at different nominal growth rates.

a CL system’s genetic construct (P_IFFL_x) co-expresses RFP and SpoTH via the AHL inducible Plux promoter. The SpoTH RBS is used as a tuning parameter (Supplementary note 5). b OL system’s genetic construct (P_OL) expresses RFP using the AHL inducible Plux promoter. c Circuit diagram illustrating how AHL induction affects ribosomes and growth rate for the open loop (OL) or closed loop (CL) systems. In the OL system, SpoTH is not present, so there is only the upper path from AHL to ribosomes. In the CL system, AHL also activates SpoTH expression and hence upregulates ribosome concentration and growth rate. Dashed edges represent sequestration of free ribosomes by a protein’s expression. d Expected growth rate as the RFP gene is activated for the ideal case, the OL, and the CL systems. e–g Growth rate normalized by the growth rate at 0 nM AHL (nominal growth rate) versus the RFP production rate for the OL and CL systems, using fructose (e), glycerol (f), and lactose (g) as the carbon source. The inset shows the nominal growth rate with no AHL induction. Data for all the RBS’s of the CL system tested, AHL induction concentrations used, and GFP production data are shown in Supplementary Fig. 2 and Supplementary Fig. 3. Data are shown as the mean ± one standard deviation (N = 3, three biological replicates). Individual experimental values are presented as gray dots. All experiments were performed in the CF945 strain. The complete experimental protocol is provided in the Materials and Methods section. Plasmid description, plasmid map, and essential DNA sequences are provided in Supplementary Information section Plasmid maps and DNA sequences.