Abstract
Lactic acid is the monomer unit of the bioplastic poly-lactic acid (PLA). One candidate organism for lactic acid production is Pichia pastoris, a yeast widely used for heterologous protein production. Nevertheless, this yeast has a poor fermentative capability that can be modulated by controlling oxygen levels. In a previous study, lactate dehydrogenase (LDH) activity was introduced into P. pastoris, enabling this yeast to produce lactic acid. The present study aimed to increase the flow of pyruvate towards the production of lactic acid in P. pastoris. To this end, a strain designated GLp was constructed by inserting the bovine lactic acid dehydrogenase gene (LDHb) concomitantly with the interruption of the gene encoding pyruvate decarboxylase (PDC). Aerobic fermentation, followed by micro-aerophilic culture two-phase fermentations, showed that the GLp strain achieved a lactic acid yield of 0.65 g/g. The distribution of fermentation products demonstrated that the acetate titer was reduced by 20% in the GLp strain with a concomitant increase in arabitol production: arabitol increased from 0.025 g/g to 0.174 g/g when compared to the GS115 strain. Taken together, the results show a significant potential for P. pastoris in producing lactic acid. Moreover, for the first time, physiological data regarding co-product formation have indicated the redox balance limitations of this yeast.
Highlights
Lactic acid has a high commercial value due to its broad application in several areas of industry, such as the automobile, food, pharmaceutical, and textile industries, in addition to the production of biodegradable polymers such as poly-lactic acid (PLA) [1]
The production of only one isomer facilitates the process of purification and polymerization into poly L-lactic acid (PLLA), which is mainly used in biomedical applications [2]
Arabitol production increased nearly seven-fold when lactate dehydrogenase (LDH) activity was associated with pyruvate decarboxylase (PDC) disruption, demonstrating that this alcohol is the main byproduct of recombinant lactic acid production in the strains of P. pastoris tested
Summary
Lactic acid has a high commercial value due to its broad application in several areas of industry, such as the automobile, food, pharmaceutical, and textile industries, in addition to the production of biodegradable polymers such as poly-lactic acid (PLA) [1]. The yeast P. pastoris has been reclassified into the new gender Komagataella, and sub-divided into the three species K. pastoris, K. phaffii and K. pseudopastoris [14] It has, as its most notable physiological feature, the ability to grow in media containing only methanol as a carbon source [15]. P. pastoris strain GS115 has an annotated gene in its genome that encodes for a putative lactate dehydrogenase (LDH) enzyme (EC 1.1.1.27). When growing it on glycerol as a substrate, lactate production is almost absent. Arabitol production increased nearly seven-fold when LDH activity was associated with PDC disruption, demonstrating that this alcohol is the main byproduct of recombinant lactic acid production in the strains of P. pastoris tested
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