Role of counterion condensation in folding of the Tetrahymena ribozyme. II. Counterion-dependence of folding kinetics.
Condensed counterions contribute to the stability of compact structures in RNA, largely by reducing electrostatic repulsion among phosphate groups. Varieties of cations induce a collapsed state in the Tetrahymena ribozyme that is readily transformed to the catalytically active structure in the presence of Mg2+. Native gel electrophoresis was used to compare the effects of the valence and size of the counterion on the kinetics of this transition. The rate of folding was found to decrease with the charge of the counterion. Transitions in monovalent ions occur 20- to 40-fold faster than transitions induced by multivalent metal ions. These results suggest that multivalent cations yield stable compact structures, which are slower to reorganize to the native conformation than those induced by monovalent ions. The folding kinetics are 12-fold faster in the presence of spermidine3+ than [Co(NH3)6]3+, consistent with less effective stabilization of long-range RNA interactions by polyamines. Under most conditions, the observed folding rate decreases with increasing counterion concentration. In saturating amounts of counterion, folding is accelerated by addition of urea. These observations indicate that reorganization of compact intermediates involves partial unfolding of the RNA. We find that folding of the ribozyme is most efficient in a mixture of monovalent salt and Mg2+. This is attributed to competition among counterions for binding to the RNA. The counterion dependence of the folding kinetics is discussed in terms of the ability of condensed ions to stabilize compact structures in RNA.
Heilman-Miller
S L
SL
Department of Chemistry, University of Maryland, College Park 20742-2021, USA.
Pan
J
J
Thirumalai
D
D
Woodson
S A
SA
D000818:Animals; D002412:Cations / Q000494:pharmacology*; D002413:Cations, Divalent / Q000494:pharmacology; D002414:Cations, Monovalent / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology; D019277:Entropy; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008958:Models, Molecular; D009690:Nucleic Acid Conformation / Q000187:drug effects*; D020871:RNA Stability / Q000187:drug effects; D016337:RNA, Catalytic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D013095:Spermidine / Q000494:pharmacology; D055672:Static Electricity; D013379:Substrate Specificity; D013768:Tetrahymena / Q000201:enzymology* / Q000235:genetics*; D014508:Urea / Q000494:pharmacology
D002412:Cations; D002413:Cations, Divalent; D002414:Cations, Monovalent; D016337:RNA, Catalytic; C032300:hexaamminecobalt(II); D003035:Cobalt; D012965:Sodium Chloride; D014508:Urea; D008274:Magnesium; D013095:Spermidine