Synthesis and Enzymic Hydrolysis of Oligoribonucleotides Incorporating 3-Deazaguanosine: The Importance of the Nitrogen-3 Atom of Single Conserved Guanosine Residues on the Catalytic Activity of the Hammerhead Ribozyme

摘要

Four base-modified hammerhead ribozyme/substrate complexes were constructed in which single guanosine (1) residues were replaced by 3-deazaguanosine (2) in the positions G_5, G_8, G_L2.1, and G_12. The base-modified ribozyme complexes were prepared by solid-phase synthesis of oligoribonucleotides employing the novel phosphoramidite 3 derived from 2. Phosphoramidite 3 carried a phenoxyacetyl group at the amino function and a diphenylcarbamoyhl residue at the oxo group of the nucleobase. The 2'-hydroxy group was blocked with a triisopropylsilyl residue. Kinetic analysis of the phosphodiester hydrolysis showed a moderate decrease of the ribozyme catalytic activity when the residues G_5 or G_8 were replaced by 3-deazaguanosine and a 200-fold decrease when G_12 was substituted. A 6-fold catalytic increase occurred when 3-deazaguanosine was replacing G_(L2.1) in the loop region. The data indicate that the N(3) atom of compound 2, in particular at position G_12 is critical for the ribozyme activity.