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Engineering Yeast for Production of Glycolic Acid

Authors: 
Koivistoinen, O., VTT Technical Research Centre of Finland
Kuivanen, J., VTT Technical Research Centre of Finland
Penttilä, M., VTT Technical Research Centre of Finland
Richard, P., VTT Technical Research Centre of Finland

Glycolic acid is the smallest α-hydroxy acid and is used in a wide range of applications e.g. cleaning, industrial detergents, cosmetics and coatings. Importantly, it can be used as a precursor for biodegradable polyglycolic acid (PGA). PGA is a strong material with an exceptionally high barrier against oxygen and is, therefore, a potential material to be applied e.g. in packaging as a barrier layer. The value chain to economically viable, bio-based PGA can potentially start from lignocellulosic carbohydrates, which after hydrolysis are converted to glycolic acid by fermentation, and further polymerized to PGA – thus providing a high-value product concept for future biorefineries.

Our approach was to test several genetically engineered yeast species to develop a fermentation based process for glycolic acid production. Glycolic acid is produced in the glyoxylate cycle by converting glyoxylate into glycolic acid by glyoxylate reductase. Yeast has an endogenous gene coding for glyoxylate reductase but in order to make the conversion more efficient, we overexpressed an NADPH-specific glyoxylate reductase from Arabidopsis thalianawith a high affinity for glyoxylate.

For efficient conversion of glyoxylate into glycolic acid, malate synthase encoding gene was deleted to prevent glyoxylate to react to malate. Additionally, steps leading to glyoxylate formation were boosted by e.g. overexpressing isocitrate lyase encoding gene. Metabolic modelling was used for analysing different pathway options and for finding the targets for further modifications to enhance glycolic acid production. The resulting strains were cultivated in shake flasks in different media and production of glycolic acid was demonstrated. The engineered Pichia kudriavzevii produced over 30 gl-1 of glycolic acid from ethanol in a fed-batch bioreactor cultivation demonstrating the technical feasibility of the process.