A Mutation in
PGM2
Causing Inefficient Galactose Metabolism in the Probiotic Yeast Saccharomyces boulardii
Authors: Liu Jing-Jing , Zhang Guo-Chang , Kong In Iok , Yun Eun Ju , Zheng Jia-Qi , Kweon Dae-Hyuk , Jin Yong-Su 
Published: March 2018

The probiotic yeast
Saccharomyces boulardii
has been extensively studied for the prevention and treatment of diarrheal diseases, and it is now commercially available in some countries.
S. boulardii
displays notable phenotypic characteristics, such as a high optimal growth temperature, high tolerance against acidic conditions, and the inability to form ascospores, which differentiate
S. boulardii
from
Saccharomyces cerevisiae
. The majority of prior studies stated that
S. boulardii
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
S. boulardii
strain, ATCC MYA-796, was examined under various culture conditions, the
S. boulardii
strain could consume galactose, but at a much lower rate than that of
S. cerevisiae
. While all
GAL
genes were present in the
S. boulardii
genome, according to analysis of genomic sequencing data in a previous study, a point mutation (G1278A) in
PGM2
, which codes for phosphoglucomutase, was identified in the genome of the
S. boulardii
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
S. cerevisiae
PGM2
in
S. boulardii
restored galactose utilization. After reverting the point mutation to a full-length
PGM2
in
S. boulardii
by Cas9-based genome editing, the growth rates of wild-type (with a truncated
PGM2
gene) and mutant (with a full-length
PGM2
) strains with glucose or galactose as the carbon source were examined. As expected, the mutant (with a full-length
PGM2
) 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
PGM2
might offer better growth on glucose at a higher temperature in return for inefficient galactose utilization. Our results suggest that the point mutation in
PGM2
might be involved in multiple phenotypes with different effects.

IMPORTANCE
Saccharomyces boulardii
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
S. boulardii
remain unknown. Our study reports that a point mutation in
PGM2
, which codes for phosphoglucomutase, is responsible for inferior galactose utilization by
S. boulardii
. After correction of the mutated
PGM2
via genome editing, the resulting strain was able to use galactose faster than a parental strain. While the
PGM2
mutation made the yeast use galactose slowly, investigation of the genomic sequencing data of other
S. boulardii
strains revealed that the
PGM2
mutation is evolutionarily conserved. Interestingly, the
PGM2
mutation was beneficial for growth at a higher temperature on glucose. We speculate that the
PGM2
mutation was enriched due to selection of
S. boulardii
in the natural habitat (sugar-rich fruits in tropical areas).