Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme.

Chowrira, Bharat M.; Berzal‐Herranz, Alfredo; Keller, C. F.; Burke, John M.

doi:10.1016/s0021-9258(19)36537-8

Cited by 76 publications

(55 citation statements)

References 39 publications

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“…By comparison, TNA has a five-atom backbone repeat unit with a four-carbon sugar that is connected by 2′,3′-phosphodiester linkages. Thus, from a purely chemical perspective, these differences are more substantial than the chemical differences separating RNA from DNA, which are incapable of undergoing transliteration even though they differ by a single 2′-hydroxyl group ( 8 , 9 ).…”

Section: Discussionmentioning

confidence: 99%

“…The extent to which different classes of nucleic acid molecules share overlapping peaks in the fitness landscape is an interesting, but largely unexplored question in molecular evolution. Classical biochemistry studies clearly show that the catalytic properties of ribozymes are not retained when the RNA sequence is prepared as DNA ( 8 , 9 ). This observation, which reflects differences in the sugar pucker and backbone inclination angle between DNA and RNA ( 10 , 11 ), led to the unwritten rule that functional activity cannot be transferred between different classes of nucleic acid molecules.…”

Section: Introductionmentioning

confidence: 99%

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Transliteration of synthetic genetic enzymes

Wang

Liu

Shehabat

et al. 2021

Nucleic Acids Research

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Functional nucleic acids lose activity when their sequence is prepared in the backbone architecture of a different genetic polymer. The only known exception to this rule is a subset of aptamers whose binding mechanism involves G-quadruplex formation. We refer to such examples as transliteration—a synthetic biology concept describing cases in which the phenotype of a nucleic acid molecule is retained when the genotype is written in a different genetic language. Here, we extend the concept of transliteration to include nucleic acid enzymes (XNAzymes) that mediate site-specific cleavage of an RNA substrate. We show that an in vitro selected 2′-fluoroarabino nucleic acid (FANA) enzyme retains catalytic activity when its sequence is prepared as α-l-threofuranosyl nucleic acid (TNA), and vice versa, a TNA enzyme that remains functional when its sequence is prepared as FANA. Structure probing with DMS supports the hypothesis that FANA and TNA enzymes having the same primary sequence can adopt similarly folded tertiary structures. These findings provide new insight into the sequence-structure-function paradigm governing biopolymer folding.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Transliteration of synthetic genetic enzymes

Wang

Liu

Shehabat

et al. 2021

Nucleic Acids Research

View full text Add to dashboard Cite

show abstract

“…Important contacts between the Tetrahymena ribozyme core and 2′-hydroxyls of both strands of the P1 helix have been identified from the loss in binding energy that results from specific deoxynucleotide substitutions (39)(40)(41)(42). Specific deoxynucleotide substitutions in loop B and the ribozyme strand of loop A also have been found to impair hairpin ribozyme cleavage activity (43,44), leading to the proposal that a "ribose zipper" motif (18) mediates docking between loops A and B (15). Functional assays developed in the current study combined with time-resolved FRET measurements of docking equilibria provide a way to evaluate energetic contributions of specific functional groups to tertiary interactions within and between hairpin ribozyme structural elements.…”

Section: Discussionmentioning

confidence: 99%

Tertiary Structure Stabilization Promotes Hairpin Ribozyme Ligation

Fedor

1999

Biochemistry

102

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The hairpin ribozyme catalyzes a reversible RNA cleavage reaction that participates in processing intermediates of viral satellite RNA replication in plants. A minimal hairpin ribozyme consists of two helix-loop-helix segments. These segments associate noncoaxially in the active folded structure in a way that brings catalytically important loop nucleotides into close proximity. The hairpin ribozyme in the satellite RNA of Tobacco Ringspot Virus assembles in the context of a four-way helical junction. Recent physical characterization of hairpin ribozyme structures using fluorescence resonance energy transfer demonstrated enhanced stability of the folded structure in the context of a four-way helical junction compared to minimal hairpin ribozyme variants. Analysis of the functional consequences of this modification of the helical junction has revealed two changes in the hairpin ribozyme kinetic mechanism. First, ribozymes with a four-way helical junction bind 3' cleavage products with much higher affinity than minimal hairpin ribozymes, evidence that tertiary interactions within the folded structure contribute to product binding energy. Second, the balance between ligation and cleavage shifts in favor of ligation. The enhanced ligation activity of hairpin ribozymes that contain a four-way helical junction supports the notion that tertiary structure stability is a major determinant of the hairpin ribozyme proficiency as a ligase and illustrates the link between RNA structure and biological function.

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“…Interestingly, the crystal structure of a group 1 ribozyme domain shows examples of functional group coordination involving both the N 3 and 2′-hydroxyl groups of adenosine (24). Since the 2′-hydroxyl of A 10 was shown to be essential for catalytic function (25), coordination involving both the N 3 and 2′-OH groups at this position cannot be ruled out as a structural motif.…”

Section: Loop 1 Ribozyme Positions a 7 -A 10mentioning

confidence: 99%

Mutational Analysis of Loops 1 and 5 of the Hairpin Ribozyme

1998

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A comprehensive analysis of base preferences for all positions in loops 1 and 5 of the hairpin ribozyme-substrate complex was carried out using a cis-ribozyme tethered to substrate by a pentapyrimidine loop. Ribozyme-substrate molecules were mutated to contain each of the three non-native base variations at each of the eight positions within these loops. Catalytic activity was measured for each mutant and compared to the activity of the original native sequence. This was the first time all base positions in these loops have been mutated to all variants and kinetically characterized. Various effects were found, ranging from invariant base positions to those with nearly complete tolerance of any base change. Two positions resulted in cleavage rates below the lower limit of accurate quantification for all non-wild-type base substitutions. These positions are G8 in the ribozyme and Gs6 in the substrate. When A10 was substituted with a pyrimidine, self-cleavage activity fell below the lower limit of detection while the remaining positions showed varying base preferences. The information reported here on loops 1 and 5 combined with previous mutagenesis data on loops 2 and 4 [Siwkowski, A., Shippy, R., and Hampel, A. (1997) Biochemistry 36, 3930-3940] completed a comprehensive mutational/kinetic analysis of every base position located within all the required loops of the hairpin ribozyme-substrate complex and allowed for the development of a mechanism for catalysis which is proposed.

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Four ribose 2'-hydroxyl groups essential for catalytic function of the hairpin ribozyme.

Cited by 76 publications

References 39 publications

Transliteration of synthetic genetic enzymes

Transliteration of synthetic genetic enzymes

Tertiary Structure Stabilization Promotes Hairpin Ribozyme Ligation

Mutational Analysis of Loops 1 and 5 of the Hairpin Ribozyme

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