Abstract
Ergothioneine (EGT) is a natural amino acid derived from histidine,
which has many active functions such as anti-oxidation,
anti-inflammation, anti-cancer and prevention of neurodegenerative
diseases. Pleurotus eryngii is a delicious edible fungus with
high EGT content. The purpose of this study was to explore the synthesis
mechanism of EGT in P. eryngii and realize heterologous expressio
of EGT in Saccharomyces cerevisiae. The EGT synthase genes in
P. eryngii were discovered by bioinformatics method, and four
synthase genes were obtained: PeEgt1, PeEgt2a,
PeEgt2b and PeEgt2c, which were heterologously expressed
in S. cerevisiae IMX581 for functional identification. Single-,
double- and triple-gene heterologous expression strains were
constructed, and the results showed that S. cerevisiae could
synthesize EGT only by integrating PeEgt1; then integrating
PeEgt2a or PeEgt2b on this basis could significantly
increase EGT production, while integrating PeEgt2c did not
increase EGT production. The yield of EGT of MX581-PeEgt1-2a-2b
was not significantly higher than that of the double-gene engineered
strains, suggesting that the low enzyme activity of PeEgt1 led to
the accumulation of S-adenosylmethionine or hercynine, which
might be the key rate-limiting step. The EGT synthase genes in P.
eryngii were mined and successfully heterologously expressed in
S. cerevisiae, and the synthesis pathway of EGT in P.
eryngii was speculated. The results of this study enrich the resource
pool of EGT synthetic genes, and provide ideas for the construction of
engineered strains with high EGT expression.