Structural Basis for a Lethal Mutation in U6 RNA
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- 作者：Dipali G. Sashital ; Anne M. Allmann ; Steven R. Van Doren ; and Samuel E. Butcher
- 刊名：Biochemistry
- 出版年：2003
- 出版时间：February 18, 2003
- 年：2003
- 卷：42
- 期：6
- 页码：1470 - 1477
- 全文大小：483K
- 年卷期：v.42,no.6(February 18, 2003)
- ISSN：1520-4995
文摘
U6 RNA is essential for nuclear pre-mRNA splicing and has been implicated directly in catalysisof intron removal. The U80G mutation at the essential magnesium binding site of the U6 3' intramolecularstem-loop region (ISL) is lethal in yeast. To further understand the structure and function of the U6 ISL,we have investigated the structural basis for the lethal U80G mutation by NMR and optical spectroscopy.The NMR structure reveals that the U80G mutation causes a structural rearrangement within the ISLresulting in the formation of a new Watson-Crick base pair (C67·G80), and disrupts a protonated C67·A79 wobble pair that forms in the wild-type structure. Despite the structural change, the accessibility ofthe metal binding site is unperturbed, and cadmium titration produces similar phosphorus chemical shiftchanges for both the U80G mutant and wild-type RNAs. The thermodynamic stability of the U80G mutantis significantly increased (
Gfold = -3.6 ± 1.9 kcal/mol), consistent with formation of the Watson-Crick pair. Our structural and thermodynamic data, in combination with previous genetic data, suggestthat the lethal basis for the U80G mutation is stem-loop hyperstabilization. This hyperstabilization mayprevent the U6 ISL melting and rearrangement necessary for association with U4.
Gfold = -3.6 ± 1.9 kcal/mol), consistent with formation of the Watson-Crick pair. Our structural and thermodynamic data, in combination with previous genetic data, suggestthat the lethal basis for the U80G mutation is stem-loop hyperstabilization. This hyperstabilization mayprevent the U6 ISL melting and rearrangement necessary for association with U4.