Genetic and biophysical characterization of GCN4 leucine zipper mutants and peptides

Abstract

The experiments presented in this thesis examined the oligomerization specificities of GCN4 mutants in vivo and association constants for formation of dimers in vitro. We have been examining the basis for oligomer choice by characterizing variants of the GCN4 leucine zipper dimerization domain containing mutations at the α positions, specifically, at α position 16. Our in vivo studies have suggested a number of interactions between wild-type GCN4 leucine zipper (containing asparagine at α position 16) and mutants with different α16 residues, and also interactions among the mutants themselves. The results of in vivo assay based on lambda repressor fusions suggest that many different zipper sequences interact and form some type of heterotypic complexes. Using sedimentation equilibrium experiments, we have correlated the phenotype determined by the repressor-based assay with dimerization of five GCN4 α position mutants, and examined the equilibrium association constants for their dimerization.