Penchovsky Robert, Breaker Ronald R
Department of Molecular, Cellular and Developmental Biology, Yale University, PO Box 208103, New Haven, Connecticut 06520-8103, USA.
Nat Biotechnol. 2005 Nov;23(11):1424-33. doi: 10.1038/nbt1155. Epub 2005 Oct 23.
Allosteric RNAs operate as molecular switches that alter folding and function in response to ligand binding. A common type of natural allosteric RNAs is the riboswitch; designer RNAs with similar properties can be created by RNA engineering. We describe a computational approach for designing allosteric ribozymes triggered by binding oligonucleotides. Four universal types of RNA switches possessing AND, OR, YES and NOT Boolean logic functions were created in modular form, which allows ligand specificity to be changed without altering the catalytic core of the ribozyme. All computationally designed allosteric ribozymes were synthesized and experimentally tested in vitro. Engineered ribozymes exhibit >1,000-fold activation, demonstrate precise ligand specificity and function in molecular circuits in which the self-cleavage product of one RNA triggers the action of a second. This engineering approach provides a rapid and inexpensive way to create allosteric RNAs for constructing complex molecular circuits, nucleic acid detection systems and gene control elements.
变构RNA作为分子开关，可根据配体结合改变折叠和功能。常见的天然变构RNA类型是核糖开关；具有相似性质的设计RNA可通过RNA工程创建。我们描述了一种用于设计由结合寡核苷酸触发的变构核酶的计算方法。创建了具有与、或、是和非布尔逻辑功能的四种通用类型的RNA开关，以模块化形式呈现，这使得在不改变核酶催化核心的情况下能够改变配体特异性。所有通过计算设计的变构核酶均被合成并在体外进行实验测试。工程化核酶表现出超过1000倍的激活，证明了精确的配体特异性，并在分子回路中发挥作用，其中一个RNA的自我切割产物触发第二个RNA的作用。这种工程方法提供了一种快速且廉价的方式来创建变构RNA，用于构建复杂的分子回路、核酸检测系统和基因控制元件。
