Ribozymes SELEX

The search for RNAs with new catalytic functions has been aided by the development of a method that rapidly searches pools of random polymers of RNA and extracts those with particular activities SELEX is nothing less than accelerated evolution in a test tube (Box 26-3). It has been used to generate RNA molecules that bind to amino acids, organic dyes, nucleotides, cyano-cobalamin, and other molecules. Researchers have isolated ribozymes that catalyze ester and amide bond formation, Sn2 reactions, metallation of (addition of metal ions to) porphyrins, and carbon-carbon bond formation. The evolution of enzymatic cofactors with nucleotide handles that facilitate their binding to ribozymes might have further expanded the repertoire of chemical processes available to primitive metabolic systems. 

Aptamers are nucleic acids which exhibit a defined structure due to their nucleotide sequence and therefore, are able to specifically bind selected targets [1] (aptus [lat.] = fitting, sticking to). Aptamers and likewise, ribozymes [2] and deoxyribozymes [3] are selected in vitro by screening nucleic acid libraries. Here we describe in detail the selection of aptamers by a process called SELEX (Systematic Evolution of Ligands by Exponential enrichment) [4]. 

The vast majority of in vitro selection experiments, however, did not focus on catalysts but on RNA molecules that spedlically bind a given target molecule, so-called aptamers. The most crucial step in any variant of this technique is selection, meaning an effective distinction between desired and undesired species. It is in the selection step that the most decisive advances in ribozyme discovery were made, as will become clear in the discussion below. The standard selection event in aptamer SELEX is a binding event, and consequently the desired molecules can be distinguished from the rest by physical separation, namely by affinity chromatography. 

In vitro selection (SELEX, Systematic Evolution of Ligands by Exponential Enrichment) has been employed for over a decade to evolve aptamers and artificial ribozymes with various catalytic functions [169, 170]. However, nucleic acids are, in contrast to proteins, limited to the four nucleobases, thus occupying a more narrow chemical space, which can limit successful SELEX experiments against particular targets. Modified nucleosides in nucleic acid libraries are useful tools in aptamer SELEX to expand the chemical space of DNA [171-174]. 

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