FIG. 5.

Band shift assays with hPvdS and mutant promoters. (A) Band shift assays were carried out as described previously (27). Core enzyme (3.4 pmol) was incubated with hPvdS (13 pmol) and digoxigenin-labeled DNA fragments containing either the pvdD wild-type promoter fragment (nucleotides −96 to +39) or a pvdD promoter fragment containing a mutation in the IS box. Following electrophoresis and transfer to a nylon membrane, the DNA-protein complexes were detected by immunoblotting with antibodies against digoxigenin. (B) Core and hPvdS were incubated with the wild-type pvdE-pvdF promoter fragment (−121/+4 with respect to the pvdE transcription start site) or with promoter fragments containing a mutation in either the pvdE IS box (mutE) or the pvdF-proximal IS box (mutF), as shown. The positions of DNA-protein complexes are indicated by arrows.