Introduction of guanidinium-modified deoxyuridine into the substrate binding regions of DNAzyme 10–23 to enhance target affinity: Implications for DNAzyme design

• Published in: Bioorganic & medicinal chemistry letters 2010-09, Vol.20 (17), p.5119-5122
• Main Authors: Lam, Curtis H., Perrin, David M.
• Format: Article
• Language: English
• Subjects: 10–23; Analytical, structural and metabolic biochemistry; Biocatalysis; Biological and medical sciences; Deoxyuridine - metabolism; DNA, Catalytic - metabolism; Dna, deoxyribonucleoproteins; DNA, Single-Stranded - metabolism; DNAzymes; Drug Design; Fundamental and applied biological sciences. Psychology; Guanidine - metabolism; Guanidinium-modified DNA; Guanidinium-modified nucleoside; Kinetics; Nucleic acids; Substrate Specificity
• ISSN: 0960-894X; 1464-3405
• DOI: 10.1016/j.bmcl.2010.07.027

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.