The genetic basis for the adaptation of E. coli to sugar synthesis from CO2

Abstract

Understanding the evolution of a new metabolic capability in full mechanistic detail is challenging, as causative mutations may be masked by non-essential "hitchhiking" mutations accumulated during the evolutionary trajectory. We have previously used adaptive laboratory evolution of a rationally engineered ancestor to generate an Escherichia coli strain able to utilize CO2 fixation for sugar synthesis. Here, we reveal the genetic basis underlying this metabolic transition. Five mutations are sufficient to enable robust growth when a non-native Calvin–Benson–Bassham cycle provides all the sugar-derived metabolic building blocks. These mutations are found either in enzymes that affect the efflux of intermediates from the autocatalytic CO2 fixation cycle toward biomass (prs, serA, and pgi), or in key regulators of carbon metabolism (crp and ppsR). Using suppressor analysis, we show that a decrease in catalytic capacity is a common feature of all mutations found in enzymes. These findings highlight the enzymatic constraints that are essential to the metabolic stability of autocatalytic cycles and are relevant to future efforts in constructing non-native carbon fixation pathways.

Point mutations in topoisomerase I alter the mutation spectrum in E. coli and impact the emergence of drug resistance genotypes

“…Biased mutation spectrum observed in two adaptive evolution experiments . We previously reported on the use of laboratory evolution in chemostats to adapt a metabolically engineered E. coli strain towards biomass synthesis from CO 2 (Antonovsky et al 2016;Herz et al 2017) . In two independent adaptive evolution experiments, we noticed a distinct bias in the types of mutation that were fixed in the population.…”

“…Tandem sequence duplication mutations are generally considered to arise from "slippage" replication events (Levinson and Gutman 1987;Zhou, Aertsen, and Michiels 2014) , however, the newly formed duplicated sequences were independent of existing sequence repeats. A detailed description regarding the adaptive laboratory evolution process and the characterization of the evolved strains has been previously reported (Antonovsky et al 2016;Herz et al 2017) .…”

Awakening a latent carbon fixation cycle inEscherichia coli

“…However, to our knowledge, the current study is the first one in which the capacity for net carbon fixation was explored in vivo using only endogenous enzymes of a heterotrophic host, thus shedding light on the emergence of novel carbon fixation pathways. Importantly, the establishment of the RuBP cycle in E. coli required long-term adaptive evolution of the microbe under selective conditions, which modulated the partitioning of metabolic fluxes between carbon fixation and biosynthetic pathways 23,63 . We expect that autotrophic growth via the GED cycle can be achieved in a similar manner.…”

Genome Editing for the Production of Natural Products in Escherichia coli

“…Laboratory‐based evolution of E. coli has been used to introduce mutations in eight different genes (prs, pgi, serA, glmU, crp, ppsR, xylA, and malT) involved in regulating fluxes between the CBB and tricarboxylic acid (TCA) cycles. However, a genetic reconstruction showed that only five of the genes are sufficient to restore the CO 2 utilizing phenotype of E. coli . Nature can therefore provide a roadmap for how to tap a carbon source that is not only abundant but is also a major industrial waste product.…”