Restoration of fitness lost due to dysregulation of the pyruvate dehydrogenase complex is triggered by ribosomal binding site modifications

Funding Information: This work was funded by the Novo Nordisk Foundation grant no. NNF10CC1016517 and National Institutes of Health grants R01GM057089 and U01AI124316. We would like to thank Marc Abrams (Systems Biology Research Group, University of California, San Diego) for his assistance with manuscript editing. A.A. and B.O.P. designed the study. A.A. C.A.O. and R.S. performed the experiments. A.A. A.V.S, A.P. L.Y. and A.M.F. analyzed the data. A.A. and B.O.P. wrote the manuscript, with contributions from all the other co-authors. The authors declare no competing interests. Publisher Copyright: © 2021 The Author(s)Pyruvate dehydrogenase complex (PDC) functions as the main determinant of the respiro-fermentative balance because it converts pyruvate to acetyl-coenzyme A (CoA), which then enters the TCA (tricarboxylic acid cycle). PDC is repressed by the pyruvate dehydrogenase complex regulator (PdhR) in Escherichia coli. The deletion of the pdhR gene compromises fitness in aerobic environments. We evolve the E. coli pdhR deletion strain to examine its achievable growth rate and the underlying adaptive strategies. We find that (1) optimal proteome allocation to PDC is critical in achieving optimal growth rate; (2) expression of PDC in evolved strains is reduced through mutations in the Shine-Dalgarno sequence; (3) rewiring of the TCA flux and increased reactive oxygen species (ROS) defense occur in the evolved strains; and (4) the evolved strains adapt to an efficient biomass yield. Together, these results show how adaptation can find alternative regulatory mechanisms for a key cellular process if the primary regulatory mode fails.Peer reviewe