Efficient Bioproduction of 3-Hydroxypropionic Acid from Methanol by a Synthetic Yeast Cell Factory

Abstract

Methanol is an ideal feedstock for bio-manufacturing chemicals without the dependence of sugars and the competition of arable lands. We here engineered an industrial yeast Pichia pastoris to efficiently produce 3-hydroxypropionic acid (3-HP) from sole methanol as a carbon source by using a malonyl-CoA-derived pathway. Optimizing the expression of malonyl-CoA reductase gene MCR from Chloroflexus aurantiacus and enhancing the supply of precursors and NADPH enabled 3-HP production of 1.5 g/L. To avoid the time-consuming genetic manipulation, metabolically transforming a free fatty acid (FFA)-overproducing strain toward 3-HP biosynthesis results in a 3-HP production of 1.9 g/L in a shake flask. Through further downregulation of methanol dissimilation, 3-HP production was improved to 2.2 g/L. Subsequent fed-batch cultivation in bioreactors achieved a remarkable 3-HP production of 48.2 g/L from minimal medium with a yield of 0.23 g/g methanol. Notably, this represents the highest reported 3-HP production from one-carbon (C1) feedstocks and is comparable to that from sugar in yeast. The high-level 3-HP production from methanol highlights the potential of P. pastoris as a workhorse for methanol biotransformation. Furthermore, the strategies presented in this study could be applied for production of other acetyl-CoA derivatives from methanol in P. pastoris.

Engineering Yeast Peroxisomes for α-Bisabolene Production from Sole Methanol with the Aid of Proteomic Analysis

“…Methanol, an important chemical feedstock, can be produced from natural gas and CO 2 by using green energy, such as solar energy . Thus, sustainable chemical production can be achieved in a carbon-neutral manner through the biotransformation of methanol, which generates high-value and complex molecules. − Pichia pastoris (also known as Komagataella phaffii) is a methylotrophic yeast that has been used to produce various proteins. , Compared with the production of valued proteins, chemical overproduction requires extensive metabolic rewiring to drive the metabolic flux toward target molecules. Although the rapid development in genetic tools facilitates the metabolic engineering of P.…”

“… 12 Thus, sustainable chemical production can be achieved in a carbon-neutral manner through the biotransformation of methanol, which generates high-value and complex molecules. 13 − 16 Pichia pastoris (also known as Komagataella phaffii ) is a methylotrophic yeast that has been used to produce various proteins. 17 , 18 Compared with the production of valued proteins, chemical overproduction requires extensive metabolic rewiring to drive the metabolic flux toward target molecules.…”

Efficient production of itaconic acid from the single-carbon substrate methanol with engineered Komagataella phaffii

“…Methylotrophic organisms, possessing the inherent capability to utilize MeOH as their exclusive carbon and energy source, stand out as promising candidates as platform organisms, with the yeast Komagataella phaffii being one of them [ 20 – 22 ]. Although K. phaffii is widely recognized in the industry for recombinant protein production [ 23 ], recent findings indicate its significant potential in generating platform chemicals [ 24 – 27 ].…”

A Light-Powered In Vitro Synthetic Enzymatic Biosystem for the Synthesis of 3-Hydroxypropionic Acid via CO2 Fixation

“…The MCR-C fragment reduces malonyl-CoA to the intermediate malonate semialdehyde (MSA), and the MCR-N fragment reduces MSA to 3-HP . Previously, Wu et al engineered an industrial yeast Pichia pastoris to efficiently produce 3-HP through CO 2 fixation using malonyl-CoA pathway . However, complex intracellular regulation is required to achieve the supply of sufficient cofactors, such as ATP and NADPH.…”