Graphical Abstract Highlights d Conversion of obligate heterotroph to full autotrophy over laboratory timescales d Non-native Calvin cycle operation generates biomass carbon from CO 2 in E. coli d Formate is oxidized by heterologous formate dehydrogenase to provide reducing power d Chemostat-based directed evolution led to complete trophic mode change in z200 days In BriefMetabolic rewiring and directed evolution led to the emergence of E. coli clones that use CO 2 as their sole carbon source, while formate is oxidized to provide all the reducing power and energy demands. SUMMARYThe living world is largely divided into autotrophs that convert CO 2 into biomass and heterotrophs that consume organic compounds. In spite of widespread interest in renewable energy storage and more sustainable food production, the engineering of industrially relevant heterotrophic model organisms to use CO 2 as their sole carbon source has so far remained an outstanding challenge. Here, we report the achievement of this transformation on laboratory timescales. We constructed and evolved Escherichia coli to produce all its biomass carbon from CO 2 . Reducing power and energy, but not carbon, are supplied via the one-carbon molecule formate, which can be produced electrochemically. Rubisco and phosphoribulokinase were co-expressed with formate dehydrogenase to enable CO 2 fixation and reduction via the Calvin-Benson-Bassham cycle. Autotrophic growth was achieved following several months of continuous laboratory evolution in a chemostat under intensifying organic carbon limitation and confirmed via isotopic labeling.

SummaryCan a heterotrophic organism be evolved to synthesize biomass from CO2 directly? So far, non-native carbon fixation in which biomass precursors are synthesized solely from CO2 has remained an elusive grand challenge. Here, we demonstrate how a combination of rational metabolic rewiring, recombinant expression, and laboratory evolution has led to the biosynthesis of sugars and other major biomass constituents by a fully functional Calvin-Benson-Bassham (CBB) cycle in E. coli. In the evolved bacteria, carbon fixation is performed via a non-native CBB cycle, while reducing power and energy are obtained by oxidizing a supplied organic compound (e.g., pyruvate). Genome sequencing reveals that mutations in flux branchpoints, connecting the non-native CBB cycle to biosynthetic pathways, are essential for this phenotype. The successful evolution of a non-native carbon fixation pathway, though not yet resulting in net carbon gain, strikingly demonstrates the capacity for rapid trophic-mode evolution of metabolism applicable to biotechnology.PaperClip

Quantitatively describing the time course of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection within an infected individual is important for understanding the current global pandemic and possible ways to combat it. Here we integrate the best current knowledge about the typical viral load of SARS-CoV-2 in bodily fluids and host tissues to estimate the total number and mass of SARS-CoV-2 virions in an infected person. We estimate that each infected person carries 109 to 1011 virions during peak infection, with a total mass in the range of 1 μg to 100 μg, which curiously implies that all SARS-CoV-2 virions currently circulating within human hosts have a collective mass of only 0.1 kg to 10 kg. We combine our estimates with the available literature on host immune response and viral mutation rates to demonstrate how antibodies markedly outnumber the spike proteins, and the genetic diversity of virions in an infected host covers all possible single nucleotide substitutions.

Many photosynthetic organisms employ a CO2 concentrating mechanism (CCM) to increase the rate of CO2 fixation via the Calvin cycle. CCMs catalyze ≈50% of global photosynthesis, yet it remains unclear which genes and proteins are required to produce this complex adaptation. We describe the construction of a functional CCM in a non-native host, achieved by expressing genes from an autotrophic bacterium in an E. coli strain engineered to depend on rubisco carboxylation for growth. Expression of 20 CCM genes enabled E. coli to grow by fixing CO2 from ambient air into biomass, with growth in ambient air depending on the components of the CCM. Bacterial CCMs are therefore genetically compact and readily transplanted, rationalizing their presence in diverse bacteria. Reconstitution enabled genetic experiments refining our understanding of the CCM, thereby laying the groundwork for deeper study and engineering of the cell biology supporting CO2 assimilation in diverse organisms.

Quantitatively describing the time course of the SARS-CoV-2 infection within an infected individual is important for understanding the current global pandemic and possible ways to combat it. Here we integrate the best current knowledge about the abundance of potential SARS-CoV-2 host cells and typical concentrations of virions in bodily fluids to estimate the total number and mass of SARS-CoV-2 virions in an infected person. We estimate that each infected person carries 109-1011 virions during peak infection, with a total mass of about 1 μg-0.1 mg, which curiously implies that all SARS-CoV-2 virions currently in the world have a mass of only 0.1-1 kg. Knowledge of the absolute number of virions in an infected individual can put into perspective parameters of the immune system response, minimal infectious doses and limits of detection in testing.