@article{delaCruzMachensMesserschmidtetal.2019,
  author    = {de la Cruz, Jorge Gonzalez and Machens, Fabian and Messerschmidt, Katrin and Bar-Even, Arren},
  title     = {Core Catalysis of the Reductive Glycine Pathway Demonstrated in Yeast},
  series = {ACS synthetic biology},
  volume    = {8},
  journal   = {ACS synthetic biology},
  number    = {5},
  publisher = {American Chemical Society},
  address   = {Washington},
  issn      = {2161-5063},
  doi       = {10.1021/acssynbio.8b00464},
  pages     = {911 -- 917},
  year      = {2019},
  abstract  = {One-carbon (C1) compounds are attractive microbial feedstocks as they can be efficiently produced from widely available resources. Formate, in particular, represents a promising growth substrate, as it can be generated from electrochemical reduction of CO2 and fed to microorganisms in a soluble form. We previously identified the synthetic reductive glycine pathway as the most efficient route for aerobic growth on formate. We further demonstrated pathway activity in Escherichia coli after expression of both native and foreign genes. Here, we explore whether the reductive glycine pathway could be established in a model microorganism using only native enzymes. We used the yeast Saccharomyces cerevisiae as host and show that overexpression of only endogenous enzymes enables glycine biosynthesis from formate and CO2 in a strain that is otherwise auxotrophic for glycine. We find the pathway to be highly active in this host, where 0.125 mM formate is sufficient to support growth. Notably, the formate-dependent growth rate of the engineered S. cerevisiae strain remained roughly constant over a very wide range of formate concentrations, 1-500 mM, indicating both high affinity for formate use and high tolerance toward elevated concentration of this C1 feedstock. Our results, as well the availability of endogenous NAD-dependent formate dehydrogenase, indicate that yeast might be an especially suitable host for engineering growth on formate.},
  language  = {en}
}
@article{HeNoorRamosParraetal.2020,
  author    = {He, Hai and Noor, Elad and Ramos-Parra, Perla A. and Garc{\´i}a-Valencia, Liliana E. and Patterson, Jenelle A. and D{\´i}az de la Garza, Roc{\´i}o I. and Hanson, Andrew D. and Bar-Even, Arren},
  title     = {In Vivo Rate of Formaldehyde Condensation with Tetrahydrofolate},
  series = {Metabolites},
  volume    = {10},
  journal   = {Metabolites},
  number    = {65},
  publisher = {MDPI},
  address   = {Basel},
  issn      = {2218-1989},
  doi       = {10.3390/metabo10020065},
  pages     = {15},
  year      = {2020},
  abstract  = {Formaldehyde is a highly reactive compound that participates in multiple spontaneous reactions, but these are mostly deleterious and damage cellular components. In contrast, the spontaneous condensation of formaldehyde with tetrahydrofolate (THF) has been proposed to contribute to the assimilation of this intermediate during growth on C1 carbon sources such as methanol. However, the in vivo rate of this condensation reaction is unknown and its possible contribution to growth remains elusive. Here, we used microbial platforms to assess the rate of this condensation in the cellular environment. We constructed Escherichia coli strains lacking the enzymes that naturally produce 5,10-methylene-THF. These strains were able to grow on minimal medium only when equipped with a sarcosine (N-methyl-glycine) oxidation pathway that sustained a high cellular concentration of formaldehyde, which spontaneously reacts with THF to produce 5,10-methylene-THF. We used flux balance analysis to derive the rate of the spontaneous condensation from the observed growth rate. According to this, we calculated that a microorganism obtaining its entire biomass via the spontaneous condensation of formaldehyde with THF would have a doubling time of more than three weeks. Hence, this spontaneous reaction is unlikely to serve as an effective route for formaldehyde assimilation.},
  language  = {en}
}