Thermodynamics of Ion-Induced RNA Folding in the Hammerhead Ribozyme: An Isothermal Titration Calorimetric Study

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• 作者：Christian Hammann ; Alan Cooper ; and David M. J. Lilley
• 刊名：Biochemistry
• 出版年：2001
• 出版时间：February 6, 2001
• 年：2001
• 卷：40
• 期：5
• 页码：1423 - 1429
• 全文大小：87K
• 年卷期：v.40,no.5(February 6, 2001)
• ISSN：1520-4995

文摘

The hammerhead ribozyme undergoes a well-defined two-stage conformational folding process,induced by the binding of magnesium ions. In this study, we have used isothermal titration calorimetryto analyze the thermodynamics of magnesium binding and magnesium ion-induced folding of the ribozyme.Binding to the natural sequence ribozyme is strongly exothermic and can be analyzed in terms of sequentialinteraction at two sites with association constants K_A = 480 and 2840 M^-1. Sequence variants of thehammerhead RNA give very different isothermal titration curves. An A14G variant that cannot undergoion-induced folding exhibits endothermic binding. By contrast, a deoxyribose G5 variant that can undergoonly the first of the two folding transitions gives a complex titration curve. However, despite thesedifferences the ITC data for all three species can be analyzed in terms of the sequential binding ofmagnesium ions at two sites. While the binding affinities are all in the region of 10³ M^-1, correspondingto free energies of G = -3.5 to -4 kcal mol^-1, the enthalpic and entropic contributions show muchgreater variation. The ITC experiments are in good agreement with earlier conformational studies of thefolding of the ion-induced folding of the hammerhead ribozyme.