Exploring energy metabolism on C1 carbon sources in the yeast Komagataella phaffii

Background.

Komagataella phaffii (Pichia pastoris) is a well-known methylotrophic yeast that can utilize methanol as the sole carbon source for growth and energy harvesting (Ata et al. 2021, Rußmayer et al. 2015). Methanol as a C1 carbon source has gained attention, as C1 carbon sources do not compete with agricultural land use and food production and can be derived from CO₂ while contributing to the mitigation of atmospheric CO₂ levels. To make an impact on the global CO₂ household, C1- based processes need to convert methanol into bulk products, such as organic acids or proteins for feed or food production.

Aims and methods.

Although K. phaffii has been extensively used for the production of recombinant proteins, its energy metabolism is still not well understood. In this PhD project, we aim to understand the contribution of the TCA cycle to energy formation in addition to the methanol dissimilation and how cytosolic NADH is supplied when the cells are grown on methanol. Understanding the energy metabolism will enable us to to understand the role of the TCA cycle during growth on methanol to be able to rewire the metabolism for an efficient bioprocess.
We will generate knockout or overexpression strains to understand which enzymes (or complexes) in TCA cycle, electron transport chain or methanol dissimilation pathway are essential and to what extent they contribute to the energy formation during the growth on methanol. The genes of the TCA cycle which are involved in the NAD⁺/NADH balance of the cells will be in the focus. Additionally, the roles of genes encoding external alternative dehydrogenases will be explored to understand how the cytosolic NADH is supplied to the respiratory chain. In addition, the interplay between methanol utilization pathway and TCA cycle for the NAD⁺/NADH balance will be investigated focusing on the NADH generating reactions.

Ata, Ö., Ergün, BG, Fickers, P. et al. What makes Komagataella phaffii non-conventional? FEMS Yeast Research 21, 8, foab059 (2021), https://doi.org/10.1093/femsyr/foab059
Rußmayer, H., Buchetics, M., Gruber, C. et al. Systems-level organization of yeast methylotrophic lifestyle. BMC Biol 13, 80 (2015). https://doi.org/10.1186/s12915-015-0186-5