Publication Date


Document Type

Doctoral Dissertation

Academic Program

Molecular Genetics and Microbiology


Molecular Genetics and Microbiology

First Thesis Advisor

Dr. Duane D. Jenness


Saccharomyces cerevisiae, Chemoreceptors


Activation mechanism of the α-factor pheromone receptor of Saccharomyces cerevisiae was analyzed using biochemical and genetic techniques. An in vitro partial proteolysis assay was developed to determine the conformational change of the receptor that occurs upon binding of agonist. The activation specific cleavages were established by comparing cleavage products with antagonist versus agonist occupied receptor. Of the changes in peptide pattern that were revealed by trypsinization, the fragment resulting from the exposure of the third loop to the protease was found to be agonist specific and to be G-protein independent. A low-affinity binding receptor mutant was isolated which failed to undergo this agonist induced conformational change. Four intra-allelic suppressors of this receptor mutant were isolated and all were mapped to the ends of transmembrane helices 4, 5, 6 and 7; all were found to be replacements of non-polar residues by polar ones. The role of the suppressor mutations in conformational change was analyzed.


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