Abstract
Engineering of nucleic acids has been a goal in research for many years. Since the discovery of catalytic nucleic acids (ribozymes and DNAzymes), this field has attracted even more attention. One reason for the increased interest is that a large number of ribozymes have been engineered that catalyze a broad range of reactions of relevance to the origin of life. Another reason is that the structures of ribozymes or DNAzymes have been modulated such that activity is dependent on allosteric regulation by an external cofactor. Such constructs have great potential for application as biosensors in medicinal or environmental diagnostics, and as molecular tools for control of cellular processes. In addition to the development of nucleic acid enzymes by in vitro selection, rational design is a powerful strategy for the engineering of ribozymes or DNAzymes with tailored features. The structures and mechanisms of a large number of nucleic acid catalysts are now well understood. Therefore, specific design of their functional properties by structural modulation is a good option for the development of custom-made molecular tools. For rational design, several parameters have to be considered, and a number of tools are available to help/guide sequence design. Here, we discuss sequence, structural and functional design using the example of hairpin ribozyme variants to highlight the challenges and opportunities of rational nucleic enzyme engineering.
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