The Dolichol Pathway Most eukaryotic membrane and secretory proteins are asparagine-linked glycoproteins. The oligosaccharide, bound to the amide N atom of an asparagine residue, influences the conformation of the protein carrying it. As a result, glycosylation affects both the physical properties of proteins and their biological functions. A group of enzymes known collectively as the dolichol pathway are responsible for the biosynthesis of the dolichol pyrophosphate-GlcNAc2Man9Glc3 precursor oligosaccharide. Dolichol pyrophosphate-GlcNAc2Man9Glc3 is then transferred to nascent polypeptides, the first step in glycoprotein biosynthesis. This early phase of glycoprotein synthesis is essential in eukaryotes and the mechanisms of dolichol pyrophosphate-GlcNAc2Man9Glc3 biosynthesis are highly conserved. The dolichol pyrophosphate-GlcNAc2Man9Glc3 is subsequently modified by the addition and removal of sugars to arrive at the staggering diversity of oligosaccharide structures observed in eukaryotes. The goal of the proposed research is to study the structures and mechanisms of the enzymes catalyzing the first seven steps of the dolichol pathway in Saccharomyces cerevisiae. Despite the importance of this pathway, very little is known about the structures and functions of the enzymes involved. Knowledge of the dolichol pathway will be valuable since a growing number of human diseases have been linked to defects in protein glycosylation. Furthermore, a thorough understanding of the mechanisms of the enzymes involved may aid efforts to synthesize specific oligosaccharides for therapeutic purposes.