ABSTRACT
Until the mid-1980s, nucleic acids were seen merely as informative molecules in opposition to proteins that carry the conformational information. This conception changed upon discovering the catalytic ability of nucleic acids (ribozymes), and especially the high afnity and specicity of short single-stranded DNA and RNA sequences (aptamers) toward a great variety of molecules, ranging from ions to whole cells.1,2 Single-stranded oligonucleotide sequences have an exceptional propensity to assume an array of secondary and tertiary structural motifs with different shapes. The number of possible thermodynamically stable structural variants of an oligonucleotide sequence is much higher than the number of variants available for a
8.1 Introduction ................................................................................................... 143 8.2 Mass Spectrometry and Chiral Discrimination ............................................. 144 8.3 Enantioselectivity Using the Chiral Recognition Ratio (CR) Method .......... 147 8.4 Deoxy Oligonucleides as Chiral Auxiliaries ................................................. 148 8.5 Amino Acid as a Co-Selector ........................................................................ 154 8.6 Relative GCA Binding Order ........................................................................ 158 8.7 DNA Quartets as Chiral Auxiliaries .............................................................. 159 8.8 Conclusion .................................................................................................... 164 Acknowledgments .................................................................................................. 164 References .............................................................................................................. 164
peptide sequence of the same length. This is simply based on the ability of nucleotide bases to interact with each other through canonical Watson-Crick as well as unusual base pairing. The existence of oligonucleotide sequences that could assume myriad shapes within a random sequence library is the basis for the remarkable success of generating aptamers to a wide variety of target molecules. Most of the aptamers reported in the literature are related to RNA sequences, but it was found that such an immobilized ligand was very quickly degraded by RNases under conventional chromatographic condition and storage. In order to overcome this severe limitation, it was important to develop an RNA molecule resistant to the classical cleaving RNases. A very interesting strategy involving the mirror-image approach was successfully developed to design biostable L-RNA ligands, also known as spiegelmers, for potential therapeutic or diagnostic applications.3 As the structure of nucleases is inherently chiral, the RNases accept only a substrate in the correct chiral conguration, i.e., the “natural” D-oligonucleotide. So, L-oligonucleotides are expected to be unsusceptible to the naturally occurring enzymes. This concept has been successfully applied to create a biostable RNA chiral stationary phase (CSP). It was demonstrated that a CSP based on L-RNA, that is, the mirror image of the natural D-RNA aptamer, was stable for an extended period of time under usual chromatographic conditions of storage and experiments.4,5 Aptamers have been used in ow cytometry,6 biosensors,7,8 afnity probe capillary electrophoresis, capillary electrochromatography, afnity chromatography, and liquid chromatography.9-15 For chiral compounds, the efcient monitoring of the selection procedure has allowed in most cases a very high specicity, exemplied by the capability of the aptamer to bind enantioselectively the target.16-19 Eric Peyrin, for the rst time, used an immobilized DNA aptamer as a new target-specic CSP for high-performance liquid chromatography.20 The enantiomers of arginine-vasopressin were separated using an immobilized 55-base DNA aptamer known to bind stereospecically the (all-D)-isomer of the oligopeptide. In a further work, such an approach has been extended to the chiral resolution of small molecules of biological interest.21 The DNA aptamers used have been selected against D-adenosine and L-tyrosinamide enantiomers. An apparent enantio separation factor of around 3.5 was observed for the anti-D-adenosine aptamer CSP, while a very high enantioselectivity was obtained with the immobilized anti-Ltyrosinamide aptamer. Structuresof aptamer complexes reveal the key molecular interactions conferringspecicity to the aptamer-ligand association, including the precisestacking of at moieties, specic hydrogen bonding, and molecularshape complementarities. These basic principles of discriminatorymolecular interactions in aptamer complexes parallel recognitionevents central to many cellular processes involving nucleic acids.
