The Thermodynamic Origin of the Stability of a Thermophilic Ribozyme

Understanding the mechanism of thermodynamic stability of an RNA structure has significant implications for the function and design of RNA. We investigated the equilibrium folding of a thermophilic ribozyme and its mesophilic homologue by using hydroxyl radical protection, small-angle x-ray scatteri...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 2001-04, Vol.98 (8), p.4355-4360
Main Authors: X.-W. Fang, Golden, B. L., Littrell, K., Shelton, V., Thiyagarajan, P., Pan, T., Sosnick, T. R.
Format: Article
Language:English
Subjects:
Base Sequence
Biochemistry
Biological Sciences
Catalysis
Catalytic activity
Circular Dichroism
Enzyme Stability
Free energy
Hydroxyl Radical
Hydroxyl radicals
Magnesium - chemistry
Melting
Molecular biology
Molecular Sequence Data
Molecular structure
Nucleotides
Protein Conformation
Proteins
Ribonucleic acid
RNA
RNA, Catalytic - chemistry
RNA, Catalytic - metabolism
Thermodynamic equilibrium
Thermodynamics
Urea - chemistry
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Summary:Understanding the mechanism of thermodynamic stability of an RNA structure has significant implications for the function and design of RNA. We investigated the equilibrium folding of a thermophilic ribozyme and its mesophilic homologue by using hydroxyl radical protection, small-angle x-ray scattering, and circular dichroism. Both RNAs require Mg2+to fold to their native structures that are very similar. The stability is measured as a function of Mg2+and urea concentrations at different temperatures. The enhanced stability of the thermophilic ribozyme primarily is derived from a tremendous increase in the amount of structure formed in the ultimate folding transition. This increase in structure formation and cooperativity arises because the penultimate and the ultimate folding transitions in the mesophilic ribozyme become linked into a single transition in the folding of the thermophilic ribozyme. Therefore, the starting point, or reference state, for the transition to the native, functional thermophilic ribozyme is significantly less structured. The shift in the reference state, and the resulting increase in folding cooperativity, is likely due to the stabilization of selected native interactions that only form in the ultimate transition. This mechanism of using a less structured intermediate and increased cooperativity to achieve higher functional stability for tertiary RNAs is fundamentally different from that commonly proposed to explain the increased stability of thermophilic proteins.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.071050698