Publisher Summary This chapter presents the methods for secreting proteins from Saccharomyces cerevisiae. The chapter focuses on methods that are used to identify and manipulate mutations affecting secretion and glycosylation in yeast. These mutations have proved useful for increasing the secreted yield of several mammalian proteins from yeast, for reducing the complexity of the carbohydrate applied to secretory giycoproteins, and for understanding the mechanism of secretion. Mutant yeast host strains have proved valuable because many mammalian secretory proteins are not secreted efficiently from wild-type yeast strains. The glycosylation pattern of proteins remaining within the cell suggests that these proteins encounter a rate-limiting step somewhere between the endoplasmic reticulum and Golgi. The protocols for screening mutagenized yeast cultures for mutants that secrete many mammalian proteins more efficiently—supersecreting mutants. These same protocols have proved useful for constructing production strains by aiding in the identification of transformants and spore progeny which secrete maximal amounts of the desired protein. Finally, these methods have been used to screen for variant proteins with greater or lesser activity than the parental form. In all of these cases, a key feature of these methods is the facility with which large numbers of yeast colonies can be screened.