A deoxyribozyme that forms a three-helix-junction complex with its RNA substrates and has general RNA branch-forming activity

Rebecca L. Coppins, Scott K Silverman

Research output: Contribution to journalArticle

Abstract

We recently used in vitro selection to identify 7S11, a deoxyribozyme that synthesizes 2′,5′-branched RNA. The 7S11 DNA enzyme mediates the nucleophilic attack of an adenosine 2′-hydroxyl group at a 5′-triphosphate, forming 2′,5′-branched RNA in a reaction that resembles the first step of in vivo RNA splicing. Here, we describe 7S11 characterization experiments that have two important implications for nucleic acid chemistry and biochemistry. First, on the basis of a comprehensive analysis of its substrate sequence requirements, 7S11 is shown to be generally applicable for the synthesis of a wide range of 2′,5′-branched RNAs. Strict substrate sequence requirements are found at the two RNA nucleotides that directly form the branched linkage, and these requirements correspond to those nucleotides found most commonly at these two positions in natural spliced RNAs. Outside of these two nucleotides, most substrate sequences are tolerated with useful ligation activity, although rates and yields vary. Because chemical synthesis approaches to branched RNA are extremely limited in scope, the deoxyribozyme-based route using 7S11 will enable many experiments that require branched RNA. Second, comprehensive nucleotide covariation experiments demonstrate that 7S11 and its RNA substrates adopt a three-helix-junction structure in which the branch-site nucleotide is located at the intersection of the three helices. Because many natural ribozymes have multi-helix junctions, 7S11 is an interesting model system for catalytic nucleic acids.

Original languageEnglish (US)
Pages (from-to)2900-2907
Number of pages8
JournalJournal of the American Chemical Society
Volume127
Issue number9
DOIs
StatePublished - Mar 9 2005

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Catalytic DNA
RNA
substrate
Nucleotides
Substrates
Nucleic acids
nucleic acid
Nucleic Acids
Catalytic RNA
Biochemistry
experiment
Experiments
biochemistry
Hydroxyl Radical
Adenosine
DNA
Enzymes
enzyme

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

Cite this

A deoxyribozyme that forms a three-helix-junction complex with its RNA substrates and has general RNA branch-forming activity. / Coppins, Rebecca L.; Silverman, Scott K.

In: Journal of the American Chemical Society, Vol. 127, No. 9, 09.03.2005, p. 2900-2907.

Research output: Contribution to journalArticle

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