Structure, expression, and regulation of UDP-GlcNAc: dolichol phosphate GlcNAc-1-phosphate transferase (DPAGT1).

Glycosylation of proteins on asparagine amino acids (N-linked) in proteins of eukaryotic cells is initiated by the biosynthesis of dolichol-pyrophosphate-N-acetylglucosamine from dolichol-phosphate and UDP-N-acetylglucosamine. The enzyme catalyzing this reaction, UDP-GlcNAc:Dolichol Phosphate GlcNAc-1-Phosphate Transferase (DPAGT1), has been further characterized in several cell types with respect to its gene, gene products, membrane topology, functional sites, lipid dependence, and metabolic regulation. This review summarizes these properties as an update from an earlier detailed and critical review by Lehrman (Lehrman, M.

Characterization of O-linked saccharides on glycoproteins.

Recombinant and native proteins of Pichia pastoris can be O-mannosylated on serine and threonine residues, allowing further elongation reactions to generate short O-linked oligosaccha-rides of mannose. Methods for release from the protein with alkaline beta-elimination with or without reduction of the released saccharides, and for subsequent chromatographic and enzymatic characterization of these saccharides are described.

Genetic engineering of Pichia pastoris to humanize N-glycosylation of proteins.

The yeast Pichia pastoris is used extensively as the host cell for large-scale production of secreted recombinant proteins. Many proteins of pharmaceutical importance are N-glycosylated, and therefore require an expression host that yields N-linked oligosaccharides that are structurally and functionally identical to the human counterpart. The recent report by Choi et al.