Production of human compatible high mannose-type (Man5GlcNAc2) sugar chains in Saccharomyces cerevisiae

A yeast mutant capable of producing Man(5)GlNAc(2) human compatible sugar chains ore glycoproteins was constructed. An expression vector for alpha-1,2-mannosidase with the "HDEL" endoplasmic reticulum retention/retrieval tag was designed and expressed in Saccharomyces cerevisiae. An in vitro alpha-1,2-mannosidase assay and Western blot analysis showed that it was successfully localized in the endoplasmic reticulum. A triple mutant yeast lacking three glycosyltransferase activities was then transformed with an alpha-1,2-mannosidase expression vector. The oligosaccharide structures of carboxypeptidase Y as well as cell surface glycoproteins were analyzed, and the recombinant yeast was shown to produce a series of high mannose-type sugar chains including Man(5)GlcNAc(2). This is the first report of a recombinant S. cerevisiae able to produce Man(5)GlcNAc(2)-oligosaccharides, the intermediate for hybrid-type and complex-type sugar chains.