Study of a hammerhead ribozyme containing 2'-modified adenosine residues

Olsen, David B.; Benseler, Fritz; Aurup, Helle; Pieken, Wolfgang A.; Eckstein, Fritz

doi:10.1021/bi00104a024

Cited by 99 publications

(55 citation statements)

References 38 publications

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“…In either case, deoxyribonucleotide substitution in the ribozyme is unexpectedly less detrimental to the catalytic core than the use of OMe nucleotides in spite of the potential conformational problem. Even so, the activity of a OMe derivative adds to the emerging notion that major parts of the catalytic core of the hammerhead ribozyme can be assembled from a variety of nucleotide building blocks [see also Pieken et al (1991)J. Relative Stability of the Analogues.…”

Section: Cedergren Unpublished Data) a Summary Of These Results Is mentioning

confidence: 99%

“…Relative Stability of the Analogues. In addition to being biochemical curiosities, oligodeoxyribonucleotide analogues such as those described above may well have in vivo applications in targeting specific cellular or viral genes (Haseloff & Gerlach, 1988;Saxver et al, 1990;Pieken et al, 1991). RNAs are well-known to be ephemeral cellular entities due to their relatively high chemical and biochemical instability.…”

Section: Cedergren Unpublished Data) a Summary Of These Results Is mentioning

confidence: 99%

“…Even though the activities reported here are less than that of the all-RNA molecule, this loss could be more than offset by the accrued stability of these mixed polymers in the primordial world. Furthermore, we cannot rule out the possibility that nucleotides other than deoxy-or ribonucleotides could furnish the desired stability without having such a pronounced effect on the activity [for example, see Pieken et al (1991)l. The ultimate question of whether DNA itself could have catalytic activity would depend on its ability to allocate other functional groups such as those on the nitrogen bases for the creation of the critical interactions which at least in the hammerhead domain are maintained by four ribonucleotides.…”

Section: Discussionmentioning

confidence: 99%

“…We have shown previously that the 2'-hydroxyl of the ribocytidine at position 17 in the substrate is essential for catalysis, a fact that corroborates the proposed cleavage mechanism involving a nucleophilic attack by this hydroxyl on the adjacent scissile phosphate bond [Perreault et al, 1990;Yang et al, 1990; see also Koizumi et al (1989)l. In contrast, no 2'-OH in the ribozyme fragment is absolutely required, even though substitution of deoxyribonucleotides at positions G5 and A9 leads to reduced catalytic activity (Perreault et al, 1991;Olsen et al, 1991). Recently, evidence for the importance of the 2'-OH of G8 has also been obtained (Williams et al, 1992).…”

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confidence: 99%

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Minimum ribonucleotide requirement for catalysis by the RNA hammerhead domain

Yang

Usman²,

Chartrand³

et al. 1992

Biochemistry

113

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Several mixed DNA/RNA and 2'-O-methylribonucleotide/RNA analogues derived from the "hammerhead" domain of RNA catalysis have been prepared to study the minimum ribonucleotide requirement for catalytic activity. Oligodeoxyribonucleotides containing from seven to as few as four ribonucleotides are active in cleaving a substrate RNA. Predominantly deoxyribonucleotide-containing analogues have kat values 20-300 and kat/& values approximately 100-2000 times lower than those of the all-RNA ribozyme. In the case of predominantly 2'Gmethyl analogues, at least five ribonucleotides are needed to assure catalytic activity. In addition, both predominantly deoxyribonucleotide and 2'Gmethyl oligomers are at least 3 orders of magnitude more stable than an all-RNA ribozyme in incubations with RNase A and a yeast extract. These results suggest that the ribophosphate backbone is not a strict requirement for ribozyme-type catalysis. The identification of the four required ribonucleotides in the hammerhead catalytic domain provides valuable information for the rational design of chemical species having ribonuclease activities.T e RNA World hypothesis, that primitive organisms made use of an RNA-based genetic system rather than the DNAbased system present in all current cell types (Gilbert, 1986), was revived by the discovery of RNA molecules, the ribozymes,

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Section: Cedergren Unpublished Data) a Summary Of These Results Is mentioning

confidence: 99%

Section: Cedergren Unpublished Data) a Summary Of These Results Is mentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Minimum ribonucleotide requirement for catalysis by the RNA hammerhead domain

Yang

Usman²,

Chartrand³

et al. 1992

Biochemistry

113

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“…1) containing either 2'-amino-2'-deoxy-or 2'-fluoro-2'-deoxynucleotides at selected positions (10,11). Substitutions by these analogues are potentially informative for two reasons.…”

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confidence: 99%

Function of specific 2'-hydroxyl groups of guanosines in a hammerhead ribozyme probed by 2' modifications.

Williams

Pieken

Eckstein

1992

Proc. Natl. Acad. Sci. U.S.A.

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The importance of the 2'-hydroxyl group of several guanosine residues for the catalytic efficiency of a hammerhead ribozyme has been investigated. Five ribozymes in which single guanosine residues were substituted with 2'-amino-, 2'-fluoro-, or 2'-deoxyguanosine were chemically synthesized. The comparison of the catalytic activity of the three 2' modifications at a specific position allows conclusions about the functional role of the parent 2'-hydroxyl group. Substitutions of nonconserved nucleotides within the ribozyme caused little alteration in the catalytic activity relative to that obtained with the unmodified ribozyme. In contrast, when either of the guanosines within the single-stranded loop between stem I and stem H of the ribozyme was replaced by 2'-deoxyguanosine or 2'-fluoro-2'-deoxyguanosine, the catalytic activities of the resulting ribozymes were reduced by factors of at least 150. The catalytic activities of the corresponding ribozymes containing 2'-amino-2'-deoxyguanosine substitutions at these positions, however, were both reduced by factors of 15. These effects resulted from decreases in the respective k,,t values, whereas variations in the Km values were comparatively small. A

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Folding of branched RNA species

Lilley

1998

Biopolymers

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The global structures of branched RNA species are important to their function. Branched RNA species are defined as molecules in which double‐helical segments are interrupted by abrupt discontinuities. These include helical junctions of different orders, and base bulges and loops. Common helical junctions are three‐ and four‐way junctions, often interrupted by mispairs or additional nucleotides. There are many interesting examples of functional RNA junctions, including the hammerhead and hairpin ribozymes, and junctions that serve as binding sites for proteins. The junctions display some common structural properties. These include a tendency to undergo pairwise helical stacking and ion‐induced conformational transitions. Helical branchpoints can act as key architectural components and as important sites for interactions with proteins. © 1999 John Wiley & Sons, Inc. Biopoly 48: 101–112, 1998

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Study of a hammerhead ribozyme containing 2'-modified adenosine residues

Cited by 99 publications

References 38 publications

Minimum ribonucleotide requirement for catalysis by the RNA hammerhead domain

Minimum ribonucleotide requirement for catalysis by the RNA hammerhead domain

Function of specific 2'-hydroxyl groups of guanosines in a hammerhead ribozyme probed by 2' modifications.

Folding of branched RNA species

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