Allosteric ribozymes sensitive to the second messengers cAMP and cGMP.
We have engineered allosteric ribozymes by combining modular rational design with combinatorial strategies. This new procedure was used to create allosteric ribozymes that are activated by specific nucleoside 3',5'-cyclic monophosphates (cNMPs). A random-sequence domain was attached to stem II of hammerhead ribozymes via a communication module that serves as an interface between ribozyme and the effector binding site. Subjecting this initial random pool to in vitro selection methods produced populations that respond, or cleave, only in the presence of specific effector molecules. From generation 18, 20 and 23, cGMP, cCMP and cAMP-specific responsive ribozymes, respectively, were isolated and characterized. These methods show great promise for engineering allosteric ribozymes and for creating new ligand-specific aptamers.
Koizumi
M
M
Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06520, USA.
Kerr
J N
JN
Soukup
G A
GA
Breaker
R R
RR
D000494:Allosteric Regulation; D020650:Combinatorial Chemistry Techniques; D000242:Cyclic AMP / Q000494:pharmacology*; D006152:Cyclic GMP / Q000494:pharmacology*; D015195:Drug Design; D012322:RNA Precursors / Q000378:metabolism*; D016337:RNA, Catalytic / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D015290:Second Messenger Systems; D014158:Transcription, Genetic
D012322:RNA Precursors; D016337:RNA, Catalytic; D000242:Cyclic AMP; D006152:Cyclic GMP