Adenine-Aptamer Complexes: A BIPARTITE RNA SITE THAT BINDS THE ADENINE NUCLEIC BASE

Marc Meli,Jacques Vergne,Jean-Luc Décout,Marie-Christine Maurel

doi:10.1074/jbc.m107130200

Abstract

RNA aptamers that are able to complex free adenine have been isolated by a SELEX (systematic evolution of ligands by exponential enrichment) procedure. The adenine binding site was revealed by sequence alignment for a prevalent cluster of aptamers, and its structure and interactions with adenine were probed by RNase digestion studies, lead cleavage, boundary determination experiments, and truncated sequences studies. A new purine binding motif was functionally and structurally characterized and compared with other RNAs specific to purine or adenylated compounds. The affinity for adenine and the specificity for other related targets were quantified. This work suggests that the adenine binding site is composed of two independent secondary structure elements forming a bipartite binding site that interacts with adenine in a new mode of purine recognition. Such binding is of great interest because the imidazole moiety is not trapped in the binding site, and would easily be available for catalytic activity.

Highlights

RNA aptamers that are able to complex free adenine have been isolated by a SELEX procedure
In vitro selection or SELEX experiments using RNA have shown that nucleic acid molecules with specific molecular recognition and functional properties can be isolated from complex pools of random sequences by repeated rounds of selection and amplification (17)
Isolation of RNA Aptamers—In vitro selection began with a pool of RNAs with a random sequence of 50 nucleotides flanked by two primer-binding regions

Summary

Introduction

RNA aptamers that are able to complex free adenine have been isolated by a SELEX (systematic evolution of ligands by exponential enrichment) procedure. Of further significance is the discovery that abasic hammerhead ribozymes can be rescued by the addition of exogenous bases that restore the catalytic activity (15) or that imidazole can rescue a cytosine mutation in a self-cleaving ribozyme (16) To this end, we have explored the possibility that regular ribonucleic acids bind adenine with the aim of forming a plausible primitive equivalent of the catalytic site of an enzyme. In vitro selection or SELEX (systematic evolution of ligands by exponential enrichment) experiments using RNA have shown that nucleic acid molecules with specific molecular recognition and functional properties can be isolated from complex pools of random sequences by repeated rounds of selection and amplification (17). Aptamers for purine compounds, ATP, theophylline, and xanthine/guanine have their own distinctive mode for purine base recognition (20 –22)

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Conclusion