[35] N-asparagine-linked oligosaccharides: Biosynthesis of the lipid-linked oligosaccharides

This chapter presents the procedures utilized by laboratories to radiolabel the dolichol-linked oligosaccharides and study their biosynthesis. Two general approaches are adopted. The chapter primarily summarizes the in vivo studies, where pulse-labeling is used to identify intermediates and determine precursor-product relationships. These studies reflect the true biosynthetic conditions and thereby allow an accurate determination of the kinetic interrelationships of the various oligosaccharide-lipid intermediates. In addition, the chapter concerns in vitro studies with isolated membrane preparations. The advantages of this approach lie in the ability to optimize label incorporated into the oligosaccharide-lipid intermediates, and the relative ease in manipulating incubation conditions to modify and test the biosynthetic machinery.