Cell lines having genetically modified glycosylation pathways that allow them
to carry out a sequence of enzymatic reactions, which mimic the processing of glycoproteins
in humans, have been developed. Recombinant proteins expressed in these engineered
hosts yield glycoproteins more similar, if not substantially identical, to their
human counterparts. Thelower eukaryotes, which ordinarily produce high-mannose
containing N-glycans, including unicellular and multicellular fungi are modified
to produce N-glycans such as Man5GlcNAc2 or other structures
along human glycosylation pathways. This is achieved using a combination of engineering
and/or selection of strains which: do not express certain enzymes which create
the undesirable complex structures characteristic of the fungal glycoproteins,
which express exogenous enzymes selected either to have optimal activity under
the conditions present in the fungi where activity is desired, or which are targeted
to an organelle where optimal activity is achieved, and combinations thereof wherein
the genetically engineered eukaryote expresses multiple exogenous enzymes required
to produce "human-like" glycoproteins.