Hydroxyl-radical footprinting using AtzR-His and the wild-type promoter region determined that the ABS consisted of three subsites. Site-directed mutagenesis of the ABS subsites combined with atzD-lacZ fusions determined that the integrity of the third ABS subsite increases the affinity of AtzR binding, and in the presence of cyanuric acid and nitrogen, the integrity of all three subsites is necessary for activation. Beta galactosidase assays confirmed the role of AtzR as an activator of atzD operon expression. EMSA confirmed binding of AtzR. comparing the mobility complexes in the presence and absence of cyanuric acid, determining a higher rate of binding in the presence
Regulated genes for each binding site are displayed below. Gene regulation diagrams show binding sites, positively-regulated genes, negatively-regulated genes, both positively and negatively regulated genes, genes with unspecified type of regulation. For each indvidual site, experimental techniques used to determine the site are also given.
|Site sequence||Regulated genes||Gene diagram||Experimental techniques||TF function||TF type|