Figure 5. Global magnesium localization along the HDV Rz’s folding pathway studied by magnesium-induced cleavage.

Different 5′-end-labeled trans-acting mutant ribozymes that halt the folding pathway at each known HDV Rz folding intermediate were folded either in the absence or the presence (+) of SdA-1 substrate or 3′-end product. The probings were then allowed to proceed for 48 h at room temperature in the presence of 50 mM Tris-HCL (pH 8.3) and 20 mM MgCl₂. A control reaction without MgCl₂ (−) was also performed. The resulting probings were analysed on 8% denaturing PAGE gels. The positions of bromophenol blue dye (BPB) and of the different regions of the Rz are indicated on the right of the gel. The lanes designated “Ladder” and “T1” represent an alkaline hydrolysis and a ribonuclease T1 (RNase T1) mapping of the wild-type version of the ribozyme, respectively. Representative guanosine residues are indicated on the left of the gel.