Figure 3.

Evaluation of the consequences of replacement of phosphate diester with a phosphorothioate internucleoside linkage in a fluorescent VS ribozyme substrate at the position designated. UM is the unmodified substrate 2. Rates of reaction (above) and % products at end point (below) were determined under pseudo first-order conditions (10 nM substrate, 75 nM ribozyme (except for G620psA621, where 375 nM ribozyme was employed), 25 mM MgCl₂, 25 mM KCl and 40 mM Tris–HCl, pH 8.0) as described in the Materials and Methods section. All data are the average of at least three independent experiments. Note that for G620psA621, the concentration of ribozyme employed to produce a measurable rate of reaction was increased from 75 nM to 375 nM, but all rate constants have been normalized with respect to ribozyme concentration to allow for this.