Building Blocks for Ribozyme Mimics:  Conjugates of Terpyridine and Bipyridine with Nucleosides

Abstract

The incorporation of the 2,2′:6′,2′′-terpyridyl (terpy) complex of Cu(II) into a deoxyoligonucleotide has led to a functional mimic of ribozymes. The resulting mimic can cleave target RNA in a sequence-directed manner by a hydrolytic mechanism. Here we describe the synthesis and characterization of four modified nucleoside phosphoramidite reagents (7, 10, 14, and 18) that contain pendant 2,2′-bipyridine or terpy ligands. The ligands are attached either to the nucleobase (7, 10) or to the sugar (14, 18). Nucleobase modification was carried out at the C-5 position of 2′-deoxyuridine (dU). One route to sugar modification was performed by synthesis of 18, a 1′functionalized analog of dU based on 1-[3-deoxy-β-D-psicofuranosyl]uracil. Another route to sugar functionaliztion resulted in 14, a 2′-O-alkyl derivative of adenosine. These modified nucleosides are building blocks for ribozyme mimics. They are designed to deliver hydrolytically active metal complexes across either the major groove (7, 10) or the minor groove (14, 18) of an RNA/DNA duplex.

Catalytic Hydrolysis of Adenosine 2‘,3‘-Cyclic Monophosphate by CuII Terpyridine

“…Despite the importance of metal-catalyzed hydrolysis of true DNA and RNA substrates, many mechanistic aspects of these reactions are unresolved. As part of our investigation of hydrolytic RNA cleavage and the design of functional mimics of ribozymes, ,,,,− we report here a detailed kinetic study of the catalytic hydrolysis of adenosine 2‘,3‘-cyclic-monophosphate (cAMP) by Cu(II) terpyridine (Cutrpy). A number of other groups have also reported functional mimics of ribozymes. ,− Here we provide evidence that a single Cutrpy interacts with one cAMP substrate in the rate-determining step of hydrolysis, and that a Cutrpy-bound hydroxide is the reactive catalyst.…”

Precise control of RNA cleavage by ribozyme mimics

“…These secondgeneration mimics function with greatly improved efficiency over our previous terpyridine reagents. 1,2 We attribute the improved cleavage to increased flexibility of the target RNA strand that results when serinol replaces a nucleotide in the DNA sequence: this eliminates a DNA/RNA base-pair near the cleavage site and may lower the barrier for phosphorane formation.…”

“…5 Our group previously reported the first wholly synthetic ribozyme mimic, 1 which was comprised of a 17-mer DNA probe with a pendant terpyridyl (terpy) complex of Cu II incorporated in DNA via a thymidine derivative. 2 Aqueous (terpy)Cu II is a known RNA transesterification and hydrolysis agent. [6][7][8] We wished to improve cleavage efficiency and the ease of preparation of ribozyme mimics.…”

The Embedded Ribonucleotide Assay: A Chimeric Substrate for Studying Cleavage of RNA by Transesterification

“…Many groups are actively developing RNA cleavage agents with possible medical applications, including the gene-specific, catalytic destruction of viral mRNA. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] Generally, catalysts are chosen or rejected based on their ability to cleave dinucleotide substrates, [20][21][22][23] although we and others have reported studies based on metal-promoted cleavage of RNA oligomers and polymers. 9,[24][25][26] The dinucleotide assays may provide misleading information about polynucleotide cleavage, due to the length-dependence of the RNA transesterification reaction (Vide infra).…”