DETECTION OF TETRAMERIZATION DOMAINS IN-VIVO BY COOPERATIVE DNA-BINDING TO TANDEM LAMBDA-OPERATOR SITES

Chimeric proteins comprising the N-terminal DNA binding domain of lamb da repressor fused to a fragment of a foreign protein have been used t o detect oligomerization of the latter. Fusions containing dimeric and tetrameric leucine zipper domains can be distinguished based on their in vivo repressor activities on a pair of cat-lacZ reporter strains. Repressor fusions are unable to efficiently repress transcription from a synthetic promoter that overlaps a weak operator site; repression b y tetrameric, but not dimeric, fusion proteins is increased by the pre sence of a strong, upstream operator site. To construct reporters we d eveloped a shuttle system that allows rapid construction of single-cop y operon fusions in E. coli, with both cat and lacZ as reporters.