Synthetic Ribozymes and Deoxyribozymes

Using the methods of in vitro selection and in vitro evolution not only the functionahty and specificity of natural ribozymes could be altered or improved, but also nucleic acid based catalysts with new catalytic activities could be evolved. The latest results in this field demonstrate that ribozymes or deoxyri- 

The isolation of new ribozymes can be achieved by either direct selection of active sequences from completely randomized pools, or by first selecting for binding to a cofactor needed in the reaction and using the aptamer as a basis for the selection of functional molecules. The more challenging method of direct selection allows a more complete search of the available sequence space, while an aptamer-based selection strategy might facilitate the isolation of functional molecules. 

Lorsch and Szostak started [5] from a specific ATP-binding RNA, which had previously been selected by affinity chromatography on ATP agarose, [4] and used it to evolve an ATP-dependent oligonucleotide kinase. [5] An RNA pool in which the central sequence of the ATP binder had been partly randomized was synthesized and was surrounded by three completely randomized sequences of a total of 100 bases (Fig. 1). This pool was incubated with ATP-y-S. RNAs onto which the y-thio-phosphate group of the ATP-y-S had been transferred could be separated from the rest 

The characterization of individual sequences from this selection experiment revealed three different structural classes of ligases from which only one, the class I ligase, generates a 3 -5 phosphodiester bond at the ligation site. [8] An optimized version of this complex ribozyme [9] that differs at 10 positions from the original sequence has a kcat of 100 min, a value comparable to that of the protein enzyme ligase (Fig. 4). [9] 

Ekland and Bartel [10] took one step further in creating a plausible scenario for an RNA world in which ribozymes catalyzing the replication of RNA are postulated. They demonstrated that the class I ribozyme is capable of extending a separate RNA primer by one nucleotide in the presence of a template oligonucleotide and nucleoside triphos- 

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Bm-gstaUer P, Famulok M (1998) Synthetic ribozymes and deoxyribozymes, in press. In Waldmann H, Mulzer J (ed) Organic Synthesis Highhghts, vol 3. Wiley-VCH, Weinheim... 

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