Group I introns are catalytic RNAs that coordinate two consecutive transesterification reactions for self-splicing. To understand how the group I intron promotes catalysis and coordinates self-splicing reactions, we determine the structures of L-16 Tetrahymena ribozyme in complex with a 5'-splice site analog product and a 3'-splice site analog substrate using cryo-EM. We solve six conformations from a single specimen, corresponding to different splicing intermediates after the first ester-transfer reaction. The structures reveal dynamics during self-splicing, including large conformational changes of the internal guide sequence and the J5/4 junction as well as subtle rearrangements of active-site metals and the hydrogen bond formed between the 2'-OH group of A261 and the N2 group of guanosine substrate. These results help complete a detailed structural and mechanistic view of this paradigmatic group I intron undergoing the second step of self-splicing.
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http://dx.doi.org/10.1038/s41467-023-36724-5 | DOI Listing |
RNA Biol
January 2024
McKetta Department of Chemical Engineering, The University of Texas at Austin, Austin, TX, USA.
Biochemistry
November 2023
Department of Molecular Biosciences, University of Texas at Austin, Austin, Texas 78712, United States.
Folding of the group I intron ribozyme and other structured RNAs has been measured using a catalytic activity assay to monitor the native state formation by cleavage of a radiolabeled oligonucleotide substrate. While highly effective, the assay has inherent limitations present in any radioactivity- and gel-based assay. Administrative and safety considerations arise from the radioisotope, and data collection is laborious due to the use of polyacrylamide gels.
View Article and Find Full Text PDFMolecules
September 2023
Graduate School of Innovative Life Science, University of Toyama, Toyama 930-8555, Toyama, Japan.
The multimolecular assembly of three-dimensionally structured proteins forms their quaternary structures, some of which have high geometric symmetry. The size and complexity of protein quaternary structures often increase in a hierarchical manner, with simpler, smaller structures serving as units for larger quaternary structures. In this study, we exploited oligomerization of a ribozyme cyclic trimer to achieve larger ribozyme-based RNA assembly.
View Article and Find Full Text PDFMol Ther Nucleic Acids
September 2023
R&D Center, Rznomics Inc, Seongnam 13486, Republic of Korea.
Circular RNA (circRNA) has various advantages over linear mRNA that is gaining success as a new vaccine and therapeutic agent. Thus, circRNA and its engineering methods have attracted attention recently. In this study, we developed a new circRNA engineering method by end-to-end self-targeting and splicing (STS) reaction using group I intron ribozyme.
View Article and Find Full Text PDFNat Struct Mol Biol
July 2023
Lady Davis Institute for Medical Research, Jewish General Hospital and Department of Anatomy and Cell Biology and Department of Medicine, McGill University, Montreal, Quebec, Canada.
Telomerase is a special reverse transcriptase ribonucleoprotein dedicated to the synthesis of telomere repeats that protect chromosome ends. Among reverse transcriptases, telomerase is unique in using a stably associated RNA with an embedded template to synthesize a specified sequence. Moreover, it is capable of iteratively copying the same template region (repeat addition processivity) through multiple rounds of RNA-DNA unpairing and reannealing, that is, the translocation reaction.
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