Ribozyme Catalysis of Metabolism in the RNA World

Chen, Xi; Li, Na; Ellington, Andrew D.

doi:10.1002/cbdv.200790055

Cited by 169 publications

(107 citation statements)

References 77 publications

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“…Dimerization was performed in 30 mM Tris-OAc (pH 7.5), 25 mM Mg(OAc) 2 , and 25 mM KOAc at 37°C for 1 h. Each sample was separated by 10% native PAGE at 4°C, and quantified with a Bio-Imaging Analyzer, BAS-2500. The gel and running buffers were identical to those used for dimerization.…”

Section: Native Polyacrylamide Gel Electrophoresismentioning

confidence: 99%

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Coordinated control of a designed trans‐acting ligase ribozyme by a loop–receptor interaction

et al. 2009

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Edited by Hans EklundKeywords: RNA ligase ribozyme DSL ribozyme Trans-acting ribozyme Modular engineering Multiple-turnover Loop-receptor interaction a b s t r a c tWe previously developed a synthetic cis-acting RNA ligase ribozyme with 3 0 -5 0 joining activity termed ''DSL'' (designed and selected ligase). DSL was easily transformed into a trans-acting form because of its highly modular architecture. In this study, we investigated the modular properties and turnover capabilities of a trans-acting DSL, tDSL-1/GUAA. tDSL-1/GUAA exhibited remarkably high activity compared with the parental cis-acting DSL, and it attained a high turnover number. Taken together, the results indicate that a loop-receptor interaction plays a significant role in determining the activity of the trans-acting ribozyme and in its ability to perform multiple turnovers of the reaction.

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Section: Native Polyacrylamide Gel Electrophoresismentioning

confidence: 99%

“…Many artificial ribozymes with a wide variety of catalytic functions have been obtained by practicing in vitro selection, demonstrating that RNA can be a versatile functional molecule [2]. Several artificial RNA ligase ribozymes have been reported [3][4][5][6][7][8].…”

Section: Introductionmentioning

confidence: 99%

Coordinated control of a designed trans‐acting ligase ribozyme by a loop–receptor interaction

et al. 2009

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“…In recent experiments, RNA has proven to have unforeseen chemical versatility (Chen et al 2007). In particular, RNA is able to specifically bind varied ligands, both fitting them to preformed binding sites and more commonly, adaptively surrounding them (Hermann and Patel 2000).…”

Section: Introductionmentioning

confidence: 99%

RNA–Amino Acid Binding: A Stereochemical Era for the Genetic Code

2009

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By combining crystallographic and NMR structural data for RNA-bound amino acids within riboswitches, aptamers, and RNPs, chemical principles governing specific RNA interaction with amino acids can be deduced. Such principles, which we summarize in a ''polar profile'', are useful in explaining newly selected specific RNA binding sites for free amino acids bearing varied side chains charged, neutral polar, aliphatic, and aromatic. Such amino acid sites can be queried for parallels to the genetic code. Using recent sequences for 337 independent binding sites directed to 8 amino acids and containing 18,551 nucleotides in all, we show a highly robust connection between amino acids and cognate coding triplets within their RNA binding sites. The apparent probability (P) that cognate triplets around these sites are unrelated to binding sites is %5.3 9 10 -45 for codons overall, and P % 2.1 9 10 -46 for cognate anticodons. Therefore, some triplets are unequivocally localized near their present amino acids. Accordingly, there was likely a stereochemical era during evolution of the genetic code, relying on chemical interactions between amino acids and the tertiary structures of RNA binding sites. Use of cognate coding triplets in RNA binding sites is nevertheless sparse, with only 21% of possible triplets appearing. Reasoning from such broad recurrent trends in our results, a majority (approximately 75%) of modern amino acids entered the code in this stereochemical era; nevertheless, a minority (approximately 21%) of modern codons and anticodons were assigned via RNA binding sites.A Direct RNA Template scheme embodying a credible early history for coded peptide synthesis is readily constructed based on these observations.

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“…in vitro evolution ͉ RNA catalysis ͉ RNA ligase ͉ selection ͉ serial transfer D arwinian evolution has been applied to populations of RNA molecules in the laboratory to generate RNA enzymes (ribozymes) that catalyze a variety of reactions (1)(2)(3). These experiments involve repeated rounds of selective amplification and mutagenesis to drive the evolving population toward desired catalytic behaviors.…”

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confidence: 99%

Emergence of a fast-reacting ribozyme that is capable of undergoing continuous evolution

Voytek

Joyce

2007

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

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It is possible to evolve RNA enzymes in a continuous manner by employing a simple serial-transfer procedure. This method was previously applied only to descendants of one unusually fastreacting RNA enzyme with RNA ligase activity. The present study establishes a second continuously evolving RNA enzyme, also with RNA ligase activity, but with a completely independent evolutionary origin. Critical to achieving the fast catalytic rate necessary for continuous evolution, development of this enzyme entailed the addition and evolutionary maturation of a 35-nucleotide accessory domain and the application of highly stringent selection pressure, with reaction times as short as 15 ms. Once established, continuous evolution was carried out for 80 successive transfers, maintaining the population against an overall dilution of 10 207 -fold. The resulting RNA enzymes exhibited Ϸ10 5 -fold improvement in catalytic efficiency, compared with the starting molecules, and became dependent on a structural feature of the substrate that previously conferred no selective advantage. This adaptation was eliminated by deleting the substrate feature and then carrying out 20 additional transfers of continuous evolution, which resulted in molecules with even greater catalytic activity. Now that two distinct species of continuously evolving enzymes have been established, it is possible to conduct molecular ecology experiments in which the two are made to compete for limited resources within a common environment.in vitro evolution ͉ RNA catalysis ͉ RNA ligase ͉ selection ͉ serial transfer D arwinian evolution has been applied to populations of RNA molecules in the laboratory to generate RNA enzymes (ribozymes) that catalyze a variety of reactions (1-3). These experiments involve repeated rounds of selective amplification and mutagenesis to drive the evolving population toward desired catalytic behaviors. Of the many ribozymes that have been obtained by in vitro evolution, six different classes catalyze the RNA-templated joining of an oligonucleotide 3Ј-hydroxyl and an oligonucleotide 5Ј-triphosphate, forming a 3Ј,5Ј-phosphodiester and releasing inorganic pyrophosphate (4-9). This reaction is similar to that carried out by protein polymerases that catalyze the synthesis of biological RNAs.All six of the known 3Ј,5Ј-ligase ribozymes achieve rate enhancements of several orders of magnitude, compared with the uncatalyzed rate of templated RNA ligation, which is Ϸ10 Ϫ7 min Ϫ1 (10). Notably, only derivatives of the class I ligase, first described by Bartel, Szostak, and colleagues (4, 11), have been able to achieve a catalytic rate of Ͼ1 min Ϫ1 . Because of the superior catalytic rate of this ribozyme, it has been chosen as the starting point for several subsequent in vitro evolution experiments. One series of studies led to the development of a sequence-general, RNA-dependent RNA polymerase ribozyme that can add as many as 20 NTPs (12, 13). This effort involved several structural modifications of the class I ligase, including the addition of a larg...

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Ribozyme Catalysis of Metabolism in the RNA World

Cited by 169 publications

References 77 publications

Coordinated control of a designed trans‐acting ligase ribozyme by a loop–receptor interaction

Coordinated control of a designed trans‐acting ligase ribozyme by a loop–receptor interaction

RNA–Amino Acid Binding: A Stereochemical Era for the Genetic Code

Emergence of a fast-reacting ribozyme that is capable of undergoing continuous evolution

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