Sequence-specific Cleavage of RNA in the Absence of Divalent Metal Ions by a DNAzyme Incorporating Imidazolyl and Amino Functionalities

Overview

Journal Nucleic Acids Res

Publisher Oxford University Press

Specialty Biochemistry

Date 2004 Mar 9

PMID 15004246

Citations 38

Authors

Alexander V Sidorov

Jane A Grasby

David M Williams

Affiliations

Soon will be listed here.

Abstract

Two modified 2'-deoxynucleoside 5'-triphosphates have been used for the in vitro selection of a modified deoxyribozyme (DNAzyme) capable of the sequence-specific cleavage of a 12 nt RNA target in the absence of divalent metal ions. The modified nucleotides, a C5-imidazolyl-modified dUTP and 3-(aminopropynyl)-7-deaza-dATP were used in place of TTP and dATP during the selection and incorporate two extra protein-like functionalities, namely, imidazolyl (histidine analogue) and primary amino (lysine analogue) into the DNAzyme. The functional groups are analogous to the catalytic Lys and His residues employed during the metal-independent cleavage of RNA by the protein enzyme RNaseA. The DNAzyme requires no divalent metal ions or other cofactors for catalysis, remains active at physiological pH and ionic strength and can recognize and cleave a 12 nt RNA substrate with sequence specificity. This is the first example of a functionalized, metal-independent DNAzyme that recognizes and cleaves an all-RNA target in a sequence-specific manner. The selected DNAzyme is two orders of magnitude more efficient in its cleavage of RNA than an unmodified DNAzyme in the absence of metal ions and represents a rate enhancement of 10(5) compared with the uncatalysed hydrolysis of RNA.

Citing Articles

Zwitterionic DNA: enzymatic synthesis of hypermodified DNA bearing four different cationic substituents at all four nucleobases.

Kuprikova N, Ondrus M, Bednarova L, Kraus T, Slavetinska L, Sykorova V Nucleic Acids Res. 2025; 53(5).

PMID: 40057376 PMC: 11890062. DOI: 10.1093/nar/gkaf155.

L-Histidine Modulates the Catalytic Activity and Conformational Changes of the HD3 Deoxyribozyme.

Sakimoto N, Imanaka H, Tomita-Sudo E, Akita T, Kawakami J Genes (Basel). 2024; 15(11).

PMID: 39596681 PMC: 11594175. DOI: 10.3390/genes15111481.

A toolbox for enzymatic modification of nucleic acids with photosensitizers for photodynamic therapy.

Niogret G, Cheriaux C, Bonhomme F, Levi-Acobas F, Figliola C, Ulrich G RSC Chem Biol. 2024; 5(9):841-852.

PMID: 39211468 PMC: 11353023. DOI: 10.1039/d4cb00103f.

In vitro evolution of ribonucleases from expanded genetic alphabets.

Jerome C, Hoshika S, Bradley K, Benner S, Biondi E Proc Natl Acad Sci U S A. 2022; 119(44):e2208261119.

PMID: 36279447 PMC: 9636917. DOI: 10.1073/pnas.2208261119.

Site-specific functionalization with amino, guanidinium, and imidazolyl groups enabling the activation of 10-23 DNAzyme.

Du S, Li Y, Chai Z, Shi W, He J RSC Adv. 2022; 10(32):19067-19075.

PMID: 35518333 PMC: 9053948. DOI: 10.1039/d0ra02226h.

References

Thum O, Jager S, Famulok M . Functionalized DNA: A New Replicable Biopolymer. Angew Chem Int Ed Engl. 2018; 40(21):3990-3993. DOI: 10.1002/1521-3773(20011105)40:21<3990::AID-ANIE3990>3.0.CO;2-O. View

Patel D, Tock M, Frary E, Feng M, Pickering T, Grasby J . A conserved tyrosine residue aids ternary complex formation, but not catalysis, in phage T5 flap endonuclease. J Mol Biol. 2002; 320(5):1025-35. DOI: 10.1016/s0022-2836(02)00547-8. View

Unwalla H, Banerjea A . Inhibition of HIV-1 gene expression by novel macrophage-tropic DNA enzymes targeted to cleave HIV-1 TAT/Rev RNA. Biochem J. 2001; 357(Pt 1):147-55. PMC: 1221937. DOI: 10.1042/0264-6021:3570147. View

Toyoda T, Imamura Y, Takaku H, Kashiwagi T, Hara K, Iwahashi J . Inhibition of influenza virus replication in cultured cells by RNA-cleaving DNA enzyme. FEBS Lett. 2000; 481(2):113-6. DOI: 10.1016/s0014-5793(00)01974-8. View

Santoro S, Joyce G, Sakthivel K, Gramatikova S, Barbas CF 3rd . RNA cleavage by a DNA enzyme with extended chemical functionality. J Am Chem Soc. 2001; 122(11):2433-9. DOI: 10.1021/ja993688s. View

Zuker M . Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 2003; 31(13):3406-15. PMC: 169194. DOI: 10.1093/nar/gkg595. View

Vaish N, Fraley A, Szostak J, McLaughlin L . Expanding the structural and functional diversity of RNA: analog uridine triphosphates as candidates for in vitro selection of nucleic acids. Nucleic Acids Res. 2000; 28(17):3316-22. PMC: 110695. DOI: 10.1093/nar/28.17.3316. View

Lee S, Sidorov A, Gourlain T, Mignet N, Thorpe S, Brazier J . Enhancing the catalytic repertoire of nucleic acids: a systematic study of linker length and rigidity. Nucleic Acids Res. 2001; 29(7):1565-73. PMC: 31265. DOI: 10.1093/nar/29.7.1565. View

Wilson D, Szostak J . In vitro selection of functional nucleic acids. Annu Rev Biochem. 2000; 68:611-47. DOI: 10.1146/annurev.biochem.68.1.611. View

10.

Cieslak M, Niewiarowska J, Nawrot M, Koziolkiewicz M, Stec W, Cierniewski C . DNAzymes to beta 1 and beta 3 mRNA down-regulate expression of the targeted integrins and inhibit endothelial cell capillary tube formation in fibrin and matrigel. J Biol Chem. 2001; 277(9):6779-87. DOI: 10.1074/jbc.M102325200. View

11.

Liu J, Lu Y . A colorimetric lead biosensor using DNAzyme-directed assembly of gold nanoparticles. J Am Chem Soc. 2003; 125(22):6642-3. DOI: 10.1021/ja034775u. View

12.

Sakthivel K, Barbas Iii C . Expanding the Potential of DNA for Binding and Catalysis: Highly Functionalized dUTP Derivatives That Are Substrates for Thermostable DNA Polymerases. Angew Chem Int Ed Engl. 2018; 37(20):2872-2875. DOI: 10.1002/(SICI)1521-3773(19981102)37:20<2872::AID-ANIE2872>3.0.CO;2-5. View

13.

Roth A, Breaker R . An amino acid as a cofactor for a catalytic polynucleotide. Proc Natl Acad Sci U S A. 1998; 95(11):6027-31. PMC: 27579. DOI: 10.1073/pnas.95.11.6027. View

14.

Li J, Zheng W, Kwon A, Lu Y . In vitro selection and characterization of a highly efficient Zn(II)-dependent RNA-cleaving deoxyribozyme. Nucleic Acids Res. 1999; 28(2):481-8. PMC: 102519. DOI: 10.1093/nar/28.2.481. View

15.

Murray J, Seyhan A, Walter N, Burke J, Scott W . The hammerhead, hairpin and VS ribozymes are catalytically proficient in monovalent cations alone. Chem Biol. 1998; 5(10):587-95. DOI: 10.1016/s1074-5521(98)90116-8. View

16.

Hampel A, Cowan J . A unique mechanism for RNA catalysis: the role of metal cofactors in hairpin ribozyme cleavage. Chem Biol. 1997; 4(7):513-7. DOI: 10.1016/s1074-5521(97)90323-9. View

17.

Nesbitt S, Hegg L, Fedor M . An unusual pH-independent and metal-ion-independent mechanism for hairpin ribozyme catalysis. Chem Biol. 1997; 4(8):619-30. DOI: 10.1016/s1074-5521(97)90247-7. View

18.

Beaudry A, Joyce G . Directed evolution of an RNA enzyme. Science. 1992; 257(5070):635-41. DOI: 10.1126/science.1496376. View

19.

Tock M, Frary E, Sayers J, Grasby J . Dynamic evidence for metal ion catalysis in the reaction mediated by a flap endonuclease. EMBO J. 2003; 22(5):995-1004. PMC: 150332. DOI: 10.1093/emboj/cdg098. View

20.

Young K, Gill F, Grasby J . Metal ions play a passive role in the hairpin ribozyme catalysed reaction. Nucleic Acids Res. 1997; 25(19):3760-6. PMC: 146958. DOI: 10.1093/nar/25.19.3760. View