Fig. 4.

Expression of CA or Ci transport improves rubisco-dependent growth of CCMB1 E. coli in intermediate CO₂ levels even in the absence of other CCM components. As shown in Fig. 3, CCMB1 E. coli strains were grown for 4 d in minimal medium in a gas-controlled plate reader. Each diamond gives the optical density (600 nm) after 4 d for one technical replicate of four biological replicates (Methods). The reference strain (“Rubisco Alone”) constitutively expresses rubisco from the p1Ac plasmid and carries a second plasmid, pFA-sfGFP, as a vector control. The reference grew only in 1.5% and 5% CO₂. The remaining strains expressed both the E. coli native β CA (can) and the DAB2 Ci transporter from a pFA-family plasmid. These two activities were isolated by means of active site mutations. The negative control “+DAB^-+CA^-” expressed inactive Can (C48A, D50A) and DAB2 (DabA2 C539A) and grew less robustly than the reference in all conditions. If either active site was left intact (“+DAB+CA^-” or “+DAB^-+CA”), we observed a sizable growth improvement in both 0.5 and 1.5% CO₂. Contrary to our expectations, this growth improvement remained when both active sites were left intact (“+DAB+CA”). Panel (B) emphasizes this effect by focusing on growth in 1.5% CO₂. See SI Appendix, Table S4 for strain genotypes and SI Appendix, Figs. S6 and S7 for growth curves and analysis of statistical significance.