Sensing complex regulatory networks by conformationally controlled hairpin ribozymes

被引:22
作者
Najafi-Shoushtari, SH [1 ]
Mayer, G [1 ]
Famulok, M [1 ]
机构
[1] Univ Bonn, Kekule Inst Organ Chem & Biochem, D-53121 Bonn, Germany
关键词
D O I
10.1093/nar/gkh643
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The hairpin ribozyme catalyses RNA cleavage by a mechanism utilizing its conformational flexibility during the docking of two independently folded internal loop domains A and B. Based on this mechanism, we designed hairpin ribozyme variants that can be induced or repressed by external effector oligonucleotides influencing the docking process. We incorporated a third domain C to assimilate alternate stable RNA motifs such as a pseudo-half-knot or an internal stem-loop structure. Small sequence changes in domain C allowed targeted switching of ribozyme activity: the same effector oligonucleotide can either serve as an inducer or repressor. The ribozymes were applied to trp leader mRNA, the RNA sequence tightly bound by l-tryptophan-activated trp-RNA-binding attenuation protein (TRAP). When domain C is complementary to this mRNA, ribozyme activity can be altered by annealing trp leader mRNA, then specifically reverted by its TRAP/tryptophan-mediated sequestration. This approach allows to precisely sense the activity status of a protein controlled by its metabolite molecule.
引用
收藏
页码:3212 / 3219
页数:8
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