A Mutation in PGM2 Causing Inefficient Galactose Metabolism in the Probiotic Yeast Saccharomyces boulardii

Abstract

The probiotic yeast has been extensively studied for the prevention and treatment of diarrheal diseases, and it is now commercially available in some countries. displays notable phenotypic characteristics, such as a high optimal growth temperature, high tolerance against acidic conditions, and the inability to form ascospores, which differentiate from The majority of prior studies stated that exhibits sluggish or halted galactose utilization. Nonetheless, the molecular mechanisms underlying inefficient galactose uptake have yet to be elucidated. When the galactose utilization of a widely used strain, ATCC MYA-796, was examined under various culture conditions, the strain could consume galactose, but at a much lower rate than that of While all genes were present in the genome, according to analysis of genomic sequencing data in a previous study, a point mutation (G1278A) in , which codes for phosphoglucomutase, was identified in the genome of the strain. As the point mutation resulted in the truncation of the Pgm2 protein, which is known to play a pivotal role in galactose utilization, we hypothesized that the truncated Pgm2 might be associated with inefficient galactose metabolism. Indeed, complementation of in restored galactose utilization. After reverting the point mutation to a full-length in by Cas9-based genome editing, the growth rates of wild-type (with a truncated gene) and mutant (with a full-length) strains with glucose or galactose as the carbon source were examined. As expected, the mutant (with a full-length ) was able to ferment galactose faster than the wild-type strain. Interestingly, the mutant showed a lower growth rate than that of the wild-type strain on glucose at 37°C. Also, the wild-type strain was enriched in the mixed culture of wild-type and mutant strains on glucose at 37°C, suggesting that the truncated might offer better growth on glucose at a higher temperature in return for inefficient galactose utilization. Our results suggest that the point mutation in might be involved in multiple phenotypes with different effects. is a probiotic yeast strain capable of preventing and treating diarrheal diseases. However, the genetics and metabolism of this yeast are largely unexplored. In particular, molecular mechanisms underlying the inefficient galactose metabolism of remain unknown. Our study reports that a point mutation in, which codes for phosphoglucomutase, is responsible for inferior galactose utilization by After correction of the mutated via genome editing, the resulting strain was able to use galactose faster than a parental strain. While the mutation made the yeast use galactose slowly, investigation of the genomic sequencing data of other strains revealed that the mutation is evolutionarily conserved. Interestingly, the mutation was beneficial for growth at a higher temperature on glucose. We speculate that the mutation was enriched due to selection of in the natural habitat (sugar-rich fruits in tropical areas).

Production of galactitol from galactose by the oleaginous yeast Rhodosporidium toruloides IFO0880

“…More likely, it is produced to prevent the intracellular accumulation of galactose-1-phosphate, a phosphorylated intermediate of the Leloir pathway (Fig. 5), which is toxic to the cell [44, 46]. The efficient utilization of galactose is limited by conversion of galactose-1-phosphate.…”

Metabolic Engineering of Saccharomyces cerevisiae to Improve Glucan Biosynthesis

“…3B). In fact, it has been reported that Pgm2 is a rate-limiting enzyme [21] and its overexpression can significantly improve galactose metabolism in S. cerevisiae [22][23][24]. Moreover, previous studies have also demonstrated that Rho1 acts as molecular switches to multiple stress conditions [25] and is involved in regulation of cell wall biogenesis in S. cerevisiae [13,14].…”

Sensitivity of yeast to lithium chloride connects the activity of YTA6 and YPR096C to translation of structured mRNAs

“…Phosphoglucomutase is responsible for converting glucose-1-phosphate to glucose-6-phosphate and LiCl is an inhibitor of its enzymatic activity. When galactose is used as the carbon source, inhibition of phosphoglucomutase by LiCl results in the accumulation of galactose metabolite intermediates that in turn causes growth defects [11,12]. In the presence of glucose, LiCl reduces the levels of UDP-glucose and disrupts the associated pathways.…”