Chatacterization of GCN4 mutants with mormal dna binding activities and decreased transcriptional activation abilities

The (R_'N4 protein, a transcriptional activator in yeast, belongs Io the AP-1 family of transcription factors and binds to the same )-1 sexluence, ATGACTCAT. us mammalian)-1 protein and Jun cncoprotein via a bZIP domain. We develotx'd a screening system which can isolate ( ;CN4 derivatives with mutations in the DNA binding domain. In orr to identify mnino acids in (FCN4 protein that plays a rnle in tJrotein-protein inmraction, we ix'r formed saturation mutagenesis in the I)NA binding)main of GCN'I using oligonueleotides containing randomized ccxton bases. The mutants were assayed for their ability to sulort transcril)tional activation, binding activity by chevking sensitivity of 3-aminotriazol, and in vitro binding assay. Two of the mutants have a normal binding activity with decreased tranrilJtional activation ability. The residues identified in these mutants may Ix: involved in the interaction with other proteins for GCN activation.