Effects of cobalt hexammine on folding and self-cleavage of the NeurosporaVS ribozyme

We have investigated the effects of Co(NH3)(6)(3+), an analog of hexahydrat ed Mg2+, on folding and catalysis of the Neurospora VS ribozyme. Most of th e metal ion-induced changes detected by chemical modification structure pro bing in either metal ion are similar, but occur at approximately 33-fold lo wer concentrations of Co(NH3)(3)(6+) than Mg2+. However, Co(NH3)(6)(3+) is not as effective at inducing two functionally important structural changes: stabilizing the pseudoknot interaction between loops I and V, and rearrang ing the secondary structure of helix Ib. Comparison of the folding of the p recursor and the downstream cleavage product, which lacks helix Ia, shows t hat helix Ia inhibits stable pseudoknot formation and rearrangement of heli x To. The VS ribozyme does not self-cleave with Co(NH3)(6)(3+) as the sole polyvalent cation; however, mixed-metal kinetic experiments show that Co(NH 3)(6)(3+) does not inhibit Mg2+- induced self-cleavage. In contrast, at sub -saturating concentrations of Mg2+, CO(NH3)(6)(3+) increases the rate of Mg 2+-induced self-cleavage, indicating that Co(NH3)(6)(3+) contributes to the functionally relevant folding of the VS ribozyme. (C) 2001 Academic Press.