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.
Highlights
IntroductionR educed one-carbon (C1) compounds are abundant in natural habitats (e.g., methanol in phyllosphere, the aerial parts of plants1) and prevalent as byproducts of industrial processes (e.g., carbon monoxide in the flue gas of the steel industry[2])
R educed one-carbon (C1) compounds are abundant in natural habitats and prevalent as byproducts of industrial processes
In a recently published paper, we demonstrated that the reductive activities of the THF enzymes and glycine cleavage/synthase system (GCS) can support the net biosynthesis of C2 and C3 compounds from formate and CO2 in E. coli.[16]
Summary
R educed one-carbon (C1) compounds are abundant in natural habitats (e.g., methanol in phyllosphere, the aerial parts of plants1) and prevalent as byproducts of industrial processes (e.g., carbon monoxide in the flue gas of the steel industry[2]). A small group of anaerobic purine- and amino-aciddegrading microbes are thought to produce glycine from one carbon units.[14,15] In a recently published paper, we demonstrated that the reductive activities of the THF enzymes and GCS can support the net biosynthesis of C2 and C3 compounds from formate and CO2 in E. coli.[16] Yet, as E. coli does not harbor an NAD-dependent formate dehydrogenase (FDH) which is vital for using formate to supply the cell with reducing power and energy it might not be an ideal host. As the enzymatic components of the reductive glycine pathway are prevalent throughout the tree of life, we wondered whether the pathway could be established using only endogenous enzymes of a model host microbe that naturally harbors NADdependent FDH. This would support the premise that C1. We further demonstrate that yeast can sustain a constant growth rate across almost 3 orders of magnitude of formate concentrations, making it an especially promising host to support the assimilation of this key C1 compound
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