DNAmoreDB - A Database of Deoxyribozymes

Published on 2004 in J. Am. Chem. Soc. volume 126 issue 50.

PubMed ID: 15600344

DOI:10.1021/ja045817x

Abstract:

We previously used in vitro selection to identify several classes of deoxyribozymes that mediate RNA ligation by attack of a hydroxyl group at a 5‘-triphosphate. In these reactions, the nucleophilic hydroxyl group is located at an internal 2‘-position of an RNA substrate, leading to 2‘,5‘-branched RNA. To obtain deoxyribozymes that instead create linear 3‘-5‘-linked (native) RNA, here we strategically modified the selection approach by embedding the nascent ligation junction within an RNA:DNA duplex region. This approach should favor formation of linear rather than branched RNA because the two RNA termini are spatially constrained by Watson−Crick base pairing during the ligation reaction. Furthermore, because native 3‘-5‘ linkages are more stable in a duplex than isomeric non-native 2‘-5‘ linkages, this strategy is predicted to favor the formation of 3‘-5‘ linkages. All of the new deoxyribozymes indeed create only linear 3‘-5‘ RNA, confirming the effectiveness of the rational design. The new deoxyribozymes ligate RNA with kobs values up to 0.5 h-1 at 37 °C and 40 mM Mg2+, pH 9.0, with up to 41% yield at 3 h incubation. They require several specific RNA nucleotides on either side of the ligation junction, which may limit their practical generality. These RNA ligase deoxyribozymes are the first that create native 3‘-5‘ RNA linkages, which to date have been highly elusive via other selection approaches.



DNAzymes linked to this article:

Name Isolated sequence Length Reaction
8AY1 CCCGACGATGGAATGGAAGGGCGGGAGAGCCGCGGTAG      38 RNA ligation
8AY3 CAGCCTACGGGTAACAAGGTGCGAGAGATCAGCGGCAG      38 RNA ligation
8AY9 CACGAGATGTTTACCACGCTGCGGGAGATTAGCGGTAG      38 RNA ligation
8AY11 CACAAGGGAACAGGCTGCGTGTGAGAGAGTCGCGGTAA      38 RNA ligation
8AY13 CAGGTCGTTAGGCGGGAGATAACAAGGTGAAGCGGTAG      38 RNA ligation
8AY17 CACGGCAACAGCAACAGCAGTGCGAGAGAGACGCGGTAG      39 RNA ligation
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