Watson-Crick pairing modification

ASON are sequences of usually 17-30 bases of single-stranded DNA that hybridize to specific genes or their mRNA products by Watson-Crick base pairing and disrupt their function. In the case of AS-ODN (antisense oligodeoxyribonucleotides) cellular RNAseH is able to bind to the DNA-RNA duplex and hydrolyze the RNA, resulting in increased transcript turnover. Modifications to the deoxy moiety at the 2 -sugar position prohibits RNAse H action. 

G and A (Fig. 12.7). Especially the 5-position of the pyrimidine DNA bases dU (and C) seems to be highly suitable for the attachment of electron donors since these base modifications can be regarded as unnatural and functionalized derivatives of the natural DNA base T. It is important to note that the design of this type of modification allows principally that the chromophores should point into the major groove of the DNA duplex while maintaining the natural Watson-Crick base pairing of the dU moiety to A as the counterbase in the complementary oligonucleotide strand. Hence this functionalization by the chromophores introduces only a local perturbation of the normal B-DNA conformation. 

This non-canonical fold, established according to chemical and enzymatic structure probing, includes an extended amino acid acceptor stem, an extra large loop instead of the T-stem and loop, and an anticodon-like domain. Hence, one or several of the six modified nucleosides are required and are responsible for its cloverleaf structure. In a further study a chimeric tRNA with the sole modification of 1-methyladenosine in position 9 was synthesized it was demonstrated that this chimeric RNA folds correctly [27]. Thus, because of Watson-Crick base-pair disruption, a single methyl group is sufficient to induce the cloverleaf folding of this unusual tRNA sequence. 

All the modifications described above for RNA interfere with Watson-Crick base pairing and thus impede the progress of reverse transcriptase... 

With many assays pushing the utility of immobilized nucleic acid components beyond simple Watson-Crick base pairing, it is not surprising that special-purpose modifications, able to add further layers of functionality, have developed alongside and been incorporated into DNA chip assays. Intrinsic qualities of the nucleic acids have been improved or modified, while others not naturally found associated with nucleic acids have been incorporated. 

Proteins, not mRNAs, are the true functional components of cells. Unlike DNA microarrays, on which interactions are based on Watson-Crick base pairing, biomo-lecular interactions on protein microarrays are determined by complex associations between the probe proteins and the target molecules. Individual protein-ligand pairs could differ greatly in their affinities. Furthermore, unlike DNA whose structure is relatively simple, proteins are extremely diverse in structure and functions, and often display many variables, such as posttranslational modifications. Protein microarrays are useful for determining numerous protein interactions including protein-protein [59], protein-DNA [26], and protein-small molecule interactions [30], or identifying the substrates of protein kinases [58]. 

Tricyclic hypermodified nucleosides are found in archaeal and eukaryotic tRNAs and are frequently observed at position 34 (wobble base) or position 37 (adjacent to the anticodon). Position 37 typically contains a hypermodified nucleoside such as N -threonylcarbamoyladenosine (t A), 2-methylthio-N -isopentenyl-ade-nosine (ms i A-37), or wybutosine (yW). yW and its derivatives occur at position 37 in archaeal and eukaryotic phenylalanine tRNA (tRNAphe). The modifications serve to maintain the correct translational reading frame via hydrophobic interactions, which reinforce codon—anticodon pairing and prevent incorrect Watson—Crick base-pairing. Studies have shown that unmodified tRNA leads to translational defects that have been implicated in different pathological states. ... 

Due to the steric constraints of Watson-Crick base pairing, modifications of bases, which do not negatively affect base pairing properties are possible only to a limited extent. Some modifications are shown in Table 9. 

Figure 8.7 Modification of oligonucleotides to increase stability, (a) Oligonucleotides (here shown as DNA) with a phosphodiester backbone (X = 0) are rapidly degraded by nucleases. Modification to create phosphorothioate analogs (X = S ) greatly increases half-life, (b) Peptide nucleic acids represent another DNA analog that can be used to bind with complementary sequences of oligonucleotides. Dashed lines represent hydrogen bonding which follows Watson-Crick base pairs.

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