Open AccessDissection of the ion-induced folding of the hammerhead ribozyme using 19 F NMR
Author(s) -
Christian Hammann,
D. Norman,
David M. J. Lilley
Publication year - 2001
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.091097498
Subject(s) - ribozyme , hairpin ribozyme , hammerhead ribozyme , vs ribozyme , chemistry , folding (dsp implementation) , mammalian cpeb3 ribozyme , ion , crystallography , rna , biochemistry , organic chemistry , electrical engineering , gene , engineering
We have used19 F NMR to analyze the metal ion-induced folding of the hammerhead ribozyme by selective incorporation of 5fluorouridine. We have studied the chemical shift and linewidths of19 F resonances of 5-fluorouridine at the 4 and 7 positions in the ribozyme core as a function of added Mg2+ . The data fit well to a simple two-state model whereby the formation of domain 1 is induced by the noncooperative binding of Mg2+ with an association constant in the range of 100 to 500 M−1 , depending on the concentration of monovalent ions present. The results are in excellent agreement with data reporting on changes in the global shape of the ribozyme. However, the NMR experiments exploit reporters located in the center of the RNA sections undergoing the folding transitions, thereby allowing the assignment of specific nucleotides to the separate stages. The results define the folding pathway at high resolution and provide a time scale for the first transition in the millisecond range.