title: Toward the combinatorial selection of chemically modified DNAzyme RNase A mimics active against all-RNA substrates.
abstract: The convenient use of SELEX and related combinatorial methods of in vitro selection provides a formidable gateway for the generation of DNA enzymes, especially in the context of improving their potential as gene therapeutic agents. Here, we report on the selection of DNAzyme 12-91, a modified nucleic acid catalyst adorned with imidazole, ammonium, and guanidinium groups that provide for efficient M(2+)-independent cleavage of an all-RNA target sequence (kobs = 0.06 min(-1)). While Dz12-91 was selected for intramolecular cleavage of an all-RNA target, it surprisingly cleaves a target containing a lone ribocytosine unit with even greater efficiency (kobs = 0.27 min(-1)) than Dz9-86 (kobs = 0.13 min(-1)). The sequence composition of Dz12-91 bears a marked resemblance to that of Dz9-86 (kobs = 0.0014 min(-1) with an all-RNA substrate) that was selected from the same library to cleave a target containing a single ribonucleotide. However, small alterations in the sequence composition have a profound impact on the substrate preference and catalytic properties. Indeed, Dz12-91 displays the highest known rate enhancement for the M(2+)-independent cleavage of all-RNA targets. Hence, Dz12-91 represents a step toward the generation of potentially therapeutically active DNAzymes and further underscores the usefulness of modified triphosphates in selection experiments.
authors.affiliation: Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver BC, V6T 1Z1, Canada.
authors.forename: Marcel
authors.initials: M
authors.lastname: Hollenstein
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Hipolito
authors.forename: Curtis H
authors.initials: CH
authors.lastname: Lam
authors.forename: David M
authors.initials: DM
authors.lastname: Perrin
mesh_terms: D001483:Base Sequence; D021881:DNA, Catalytic / Q000737:chemistry* / Q000378:metabolism*; D012313:RNA / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000378:metabolism*; D052156:SELEX Aptamer Technique; D013379:Substrate Specificity
chemical_list: D021881:DNA, Catalytic; D012313:RNA; D012259:Ribonuclease, Pancreatic