Recoding UAG to selenocysteine in Saccharomyces cerevisiae

Area of Science:

• Biochemistry
• Molecular Biology
• Synthetic Biology

Background:

• Selenocysteine (Sec) incorporation imparts unique properties to proteins.
• Eukaryotic selenoprotein production is hindered by the loss of the Sec biosynthesis pathway in fungi.
• Previous success in bacterial Sec production informed this study.

Purpose of the Study:

• To establish a functional Sec biosynthesis pathway in Saccharomyces cerevisiae.
• To enable site-specific genetic encoding of Sec in yeast.
• To facilitate recombinant production of eukaryotic selenoproteins.

Main Methods:

• Designed a novel Sec biosynthesis pathway in yeast using bacterial components.
• Mutated yeast tRNASer to mimic bacterial tRNASec for recognition by specific enzymes.
• Combined pathway engineering with metabolic engineering for Sec incorporation.

Main Results:

• Successfully established a novel Sec biosynthesis pathway in Saccharomyces cerevisiae.
• Demonstrated site-specific incorporation of Sec into a target protein.
• Produced active methionine sulfate reductase containing genetically encoded Sec.

Conclusions:

• Yeast can be engineered for selenoprotein production through site-specific Sec incorporation.
• This work overcomes evolutionary loss of the Sec pathway in fungi.
• Opens avenues for facile recombinant production of valuable selenoproteins in yeast.