Examination of the catalytic fitness of the hammerhead ribozyme by in vitro selection.

摘要: We have designed a self-cleaving ribozyme construct that is rendered inactive during preparative in vitro transcription by allosteric interactions with ATP. This was constructed joining hammerhead domain to an ATP-binding RNA aptamer, thereby creating whose catalytic rate can be controlled Upon purification PAGE, the engineered undergoes rapid self-cleavage when incubated absence of strategy "allosteric delay" used prepare intact ribozymes would otherwise self-destruct transcription. Using similar strategy, we prepared combinatorial pool order assess fitness carry natural consensus sequence for hammerhead. selection, this comprehensive screened variants also display activity. find sequences comprise core naturally occurring dominate population selected RNAs, indicating optimal function.