Structure-function studies of the Saccharomyces cerevisiae a-factor lipopeptide mating pheromone

Mating in Saccharomyces cerevisiae is initiated by extracellular peptides, known as mating pheromones. The mating factors are recognized by membrane-bound receptors which are similar to receptors in mammalian cells. The pheromone system of the S. cerevisiae can be used as a model to understand the mode of action of peptide hormones in more complex mammalian systems. The purpose of this thesis is to provide information regarding the yeast lipopeptide pheromone, a-factor. The first part of this project, described in Chapter 2, was the development of an assay which could be used to define the binding of a-factor to its receptor. The assay, which made use of membrane preparations in place of whole cells, was able to show that a larger percentage of the radioactive a-factor pheromone did bind to membranes which contained the a-factor receptor (Ste3p) when compared to membranes which lacked the receptor. However, further experiments are needed to more clearly define the pheromone/receptor interaction. The second part of this project, described in Chapter 3, was the examination of the role of the farnesyl group in a-factor bioactivity, particularly mating restoration. This examination made use of a-factors with altered farnesyl groups that contained the novel lipid substitutions of the 3-methyl moiety with bulkier additions. The study suggested that there is a specific preference for a particular relationship between the membrane and the ligand with regard to bioactivity, and that this preference is mediated via the farnesyl group. Thus, the farnesyl group may also play a role in presentation of the peptide portion to the receptor in addition to its role in membrane localization.