FIG. 5.

DNase I footprinting analysis of N. meningitidis Fur with the nspA operator probe. (A) Footprinting analysis. The plus strand of the nspA promoter fragment was labeled and incubated with increasing concentrations of N. meningitidis Fur prior to digestion with DNase I. Lane 1, no Fur; lane 2, 150 nM Fur; lane 3, 460 nM Fur; lane 4, 1.22 μM Fur. DNA standards (GATC) are shown on the right. The region protected by Fur is indicated by the dark line. (B) Schematic representation of the operator elements of the iron-activated nspA, aniA, and secY genes and the iron-repressed fur gene. DNA sequences of the iron-activated and Fur-regulated nspA and aniA genes, the iron-repressed and Fur-regulated fur gene, and the iron-activated secY gene operators were amplified for in vitro binding studies using the primers listed in Table 1. Boxed nucleotides represent the in silico-identified Fur box. The previously reported hexameric repeat 5′-NATW(A/T)AT-3′ (17) is represented in boldface type. In the case of the nspA gene, the extent of the footprinted region is represented by a double-headed arrow.