Guo Feng, Cech Thomas R
Howard Hughes Medical Institute, Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309-0215, USA.
Nat Struct Biol. 2002 Nov;9(11):855-61. doi: 10.1038/nsb850.
Determining how large RNA molecules stabilize their tertiary structures is critical for understanding how they perform their biological functions. Here we use in vitro selection to identify active variants of the Tetrahymena ribozyme with increased stability. The mutant pool converged to a single family that shared nine mutations; an RNA representing the consensus sequence was structurally more stable by 10.5 degrees C and catalytically active at elevated temperatures. Remarkably, of the nine altered sites, most are already known to be involved in tertiary interactions, and the stabilizing mutations primarily improve the packing interactions in the molecular interior. The wild type ribozyme and the selected mutants provide pairs of mesophilic and thermophilic homologs for studying the origin of their thermal stability.
确定RNA大分子如何稳定其三级结构对于理解它们如何执行生物学功能至关重要。在这里，我们使用体外筛选来鉴定具有更高稳定性的嗜热四膜虫核酶的活性变体。突变体库汇聚到一个共有九个突变的单一家族；代表共有序列的RNA在结构上更稳定，提高了10.5摄氏度，并且在升高的温度下具有催化活性。值得注意的是，在这九个改变的位点中，大多数已知已经参与三级相互作用，并且稳定突变主要改善了分子内部的堆积相互作用。野生型核酶和选定的突变体为研究它们热稳定性的起源提供了嗜温和嗜热同源物对。
