An aminoacylation ribozyme evolved from a natural tRNA-sensing T-box riboswitch

Ishida, Satoshi; Terasaka, Naohiro; Katoh, Takayuki; Suga, Hiroaki

doi:10.1038/s41589-020-0500-6

Cited by 30 publications

(39 citation statements)

References 48 publications

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“…In recent years, synthetic biologists have started to create synthetic tRNA-aminoacylation protein enzymes and ribozymes [ 18 , 72 , 73 ]. However, unlike most of the natural aaRS species, these artificial protein enzymes, including my HisRS chimeras, are composed of separate domains fused with a flexible linker [ 18 , 72 ].…”

Section: Discussionmentioning

confidence: 99%

“…Thus, such artificial enzymes were far less active and precise than natural enzymes [ 18 , 72 ]. On the other hand, the ribozymes required pre-activated amino acid substrates rather than free amino acid and ATP [ 73 ]. An apparent drawback of my allo-tRNA strategy is that the tip of the V-arm is distant from the CCA tail of tRNA.…”

Section: Discussionmentioning

confidence: 99%

“…It may be helpful to use a long and rigid linker motif found in mitochondrial AlaRS species [ 74 ] or to use a long-D-arm binding protein domain to wrap around tRNA [ 37 , 38 ]. The use of T-box riboswitches as RNA scaffolds may be an alternative approach [ 73 , 75 ]. In summary, the novel allo-tRNA chassis, or orthogonal allo-tRNAs, would facilitate creation of synthetic tRNA-aminoacylation enzymes and ribozymes.…”

Section: Discussionmentioning

confidence: 99%

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Rational Design of Aptamer-Tagged tRNAs

Mukai

2020

IJMS

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Reprogramming of the genetic code system is limited by the difficulty in creating new tRNA structures. Here, I developed translationally active tRNA variants tagged with a small hairpin RNA aptamer, using Escherichia coli reporter assay systems. As the tRNA chassis for engineering, I employed amber suppressor variants of allo-tRNAs having the 9/3 composition of the 12-base pair amino-acid acceptor branch as well as a long variable arm (V-arm). Although their V-arm is a strong binding site for seryl-tRNA synthetase (SerRS), insertion of a bulge nucleotide in the V-arm stem region prevented allo-tRNA molecules from being charged by SerRS with serine. The SerRS-rejecting allo-tRNA chassis were engineered to have another amino-acid identity of either alanine, tyrosine, or histidine. The tip of the V-arms was replaced with diverse hairpin RNA aptamers, which were recognized by their cognate proteins expressed in E. coli. A high-affinity interaction led to the sequestration of allo-tRNA molecules, while a moderate-affinity aptamer moiety recruited histidyl-tRNA synthetase variants fused with the cognate protein domain. The new design principle for tRNA-aptamer fusions will enhance radical and dynamic manipulation of the genetic code.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Rational Design of Aptamer-Tagged tRNAs

Mukai

2020

IJMS

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“…A recent study delineates such a pathway for the evolution of an aminoacylation ribozyme which could be part of a primitive translation system. 123 An aminoacylation ribozyme evolved from a natural tRNAsensing T-box riboswitch. The naturally occurring T-box riboswitches selectively sense the aminoacylation status of cognate tRNAs.…”

Section: Riboswitch Goes Ribozyme -And An Outlook On Non-protein Rna mentioning

confidence: 99%

Fundamental studies of functional nucleic acids: aptamers, riboswitches, ribozymes and DNAzymes

2020

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“…The intricate and specific interactions between the T-box:tRNA pair can be leveraged for a variety of applications across basic research and bioengineering. For example, a recent study used a glyQS T-box to engineer a ribozyme that can specifically charge tRNA gly for use in cell-free protein synthesis [19]. Tboxes also have the potential to be used as a generalizable 'registry-of-parts', capable of independently sensing amino acid levels in the environment with a single RNA element [20].…”

Section: Introductionmentioning

confidence: 99%

TBDB – A database of structurally annotated T-box riboswitch:tRNA pairs

Marchand

Smela

Narasimhan

et al. 2020

Preprint

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AbstractT-box riboswitches constitute a large family of tRNA-binding leader sequences that play a central role in gene regulation in many gram-positive bacteria. Accurate inference of the tRNA binding to T-boxes is critical to predict their cis-regulatory activity. However, there is no central repository of information on the tRNA binding specificities of T-box riboswitches and de novo prediction of binding specificities requires advance knowledge of computational tools to annotate riboswitch secondary structure features. Here we present T-box annotation Database (TBDB,https://tbdb.io), an open-access database with a collection of 23,497 T-box sequences, spanning the major phyla of 3,621 bacterial species. Among structural predictions, the TBDB also identifies specifier sequences, cognate tRNA binding partners, and downstream regulatory target. To our knowledge, the TBDB presents the largest collection of feature, sequence, and structural annotations carried out on this important family of regulatory RNA.

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An aminoacylation ribozyme evolved from a natural tRNA-sensing T-box riboswitch

Cited by 30 publications

References 48 publications

Rational Design of Aptamer-Tagged tRNAs

Rational Design of Aptamer-Tagged tRNAs

Fundamental studies of functional nucleic acids: aptamers, riboswitches, ribozymes and DNAzymes

TBDB – A database of structurally annotated T-box riboswitch:tRNA pairs

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