Replication data for "'Metabolic engineering of Komagataella phaffii for enhanced 3-hydroxypropionic acid production from methanol"

Additional files of research article "Metabolic engineering of Komagataella phaffii for enhanced 3-hydroxypropionic acid (3-HP) production from methanol". This research paper describes a combinatorial metabolic engineering strategy aiming at increasing the supply of metabolic precursors and 3-HP export, applied to the original 3-HP producing K. phaffii strain harboring the synthetic β-alanine pathway and a heterologous NADP-dependent formate dehydrogenase.

Several strains overexpressing the genes encoding enzymes catalyzing reactions immediately upstream of the β-alanine pathway were constructed. To further channel flux through the heterologous NADP-dependent formate dehydrogenase, we deleted the endogenous NAD-dependent analogous enzyme. Finally, we sought to promote 3-HP export by overexpressing two monocarboxylate permeases. The constructed strains were tested on 24 deep-well plate cultivations containing buffered minimal medium (BMM) with methanol. Three independent transformants from each strain were tested. Final biomass concentration as well as methanol and 3-HP quantification were determined at the end of the cultures.

The best-performing strains were further characterized in aerobic fed-batch reactors following a pre-programmed exponential feeding strategy for controlled growth rate at pH 5 and 3.5. Offline and online state variables during the fed-batch phase of the bioreactor-scale experiments were monitored and recorded. Experiments at pH 5 were conducted in duplicate, while a single cultivation replicate for each strain was performed at pH 3.5.

The files attached herein contain: a description of the molecular cloning materials and methods, raw data obtained from the 24 deep-well plate cultivations, raw data of the online monitored standard process parameters, and raw and processed data obtained from the bioreactor-scale experiments.