Introduction of guanidinium-modified deoxyuridine into the substrate binding regions of DNAzyme 10–23 to enhance target affinity: Implications for DNAzyme design
✍ Scribed by Curtis H. Lam; David M. Perrin
- Book ID
- 104004639
- Publisher
- Elsevier Science
- Year
- 2010
- Tongue
- English
- Weight
- 355 KB
- Volume
- 20
- Category
- Article
- ISSN
- 0960-894X
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✦ Synopsis
Deoxyribozymes (DNAzymes) are important catalysts for potential therapeutic RNA destruction and no DNAzyme has received as much notoriety in terms of therapeutic use as the Mg(2+)-dependent RNA-cleaving DNAzyme 10-23 (Dz10-23). As such, we have investigated the synthetic modification of Dz10-23 with a guanidinium group, a functionality that reduces the anionic nature and can potentially enhance the membrane permeability of oligonucleotides. To accomplish this, we synthesized a heretofore unknown phosphoramidite, 5-(N,N'-biscyanoethoxycarbonyl)-guanidinoallyl-2'-deoxyuridine and then incorporated it into oligonucleotides via solid phase synthesis to study duplex stability and its effect on Dz10-23. This particular modification was chosen as it had been used in the selection of Mg(2+)-free self-cleaving DNAzymes; as such this will enable the eventual comparison of modified DNAzymes that do or do not depend on Mg(2+) for catalysis. Consistent with antecedent studies that have incorporated guanidinium groups into DNA oligonucleotides, this guanidinium-modified deoxyuridine enhanced the thermal stability of resulting duplexes. Surprisingly however, Dz10-23, when synthesized with modified residues in the substrate binding regions, was found to be somewhat less active than its non-modified counterpart. This work suggests that this particular system exhibits uniform binding with respect to ground state and transition state and provides insight into the challenge of re-engineering a Mg(2+)-dependent DNAzyme with enhanced catalytic activity.