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Separate metal requirements for loop interactions and catalysis in the extended hammerhead ribozyme
Nak-Kyoon Kim1, Ayaluru Murali, Victoria J DeRose
1Department of Chemistry, Texas A&M University, College Station, TX 77843, USA.
Abstract:
An extended hammerhead ribozyme derived from Schistosoma mansoni, including conserved loops in stems I and II, has been examined to directly monitor the relationship between docking of loops and its activity using site-directed spin labeling (SDSL) and EPR spectroscopy. Dynamics with EPR spectroscopy and fast-quench kinetics measurements have shown that the docking of stems I and II occurs at low Mg2+ concentrations ([Mg2+]1/2,dock = 0.7 mM, 0.1 M NaCl), but a much weaker Mg2+ interaction ([Mg2+]1/2,cat approximately 90 mM) increases activity to very high maximum rates of approximately 1 s-1 at 0.1 M Na+ and pH 7.0.
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