FIG. 4.

Primer extension analysis of the promoters of the frpB-katA and ceuE-tsaA loci. Total RNAs isolated from H. pylori strains G27 (lanes 1 and 2) and G27fur::kan (lanes 3 and 4) were hybridized to the radiolabeled oligonucleotides Frp (CTCTTAAAAACATCCAAC) (a), Kat (CACATCTTTATTAACCAT) (b), Ceu (ACGATGAAACAAGAAGCG) (c), and Tsa (GGCAAGTTTTGTAACTAAC) (d) and elongated with reverse transcriptase (27). Elongated primers are indicated by arrows. To ensure correct mapping of the extension, we sequenced in parallel the respective cloned promoter region with the same primers used in the primer extension reactions (not shown). The nucleotide sequence of the sense strand upstream of the transcriptional initiations are shown to the left of the panels, with the −10 motifs indicated by a vertical bar and the nucleotides corresponding to +1 initiation sites indicated by bent arrows. Wild-type H. pylori G27 and the isogenic fur mutant G27fur:: kan were grown to logarithmic phase in liquid cultures and then harvested or further incubated for 15 min in the presence of 50 μM 2,2′-dipyridyl before being harvested (lanes 2 and 4, marked by −).