Dataset Information

Mimicking Co-Transcriptional RNA Folding Using a Superhelicase.

ABSTRACT: Vectorial folding of RNA during transcription can produce intermediates with distinct biochemical activities. Here, we design an artificial minimal system to mimic cotranscriptional RNA folding in vitro. In this system, a presynthesized RNA molecule begins to fold from its 5'-end, as it is released from a heteroduplex by an engineered helicase that translocates on the complementary DNA strand in the 3'-to-5' direction. This chemically stabilized "superhelicase" Rep-X processively unwinds thousands of base pairs of DNA. The presynthesized RNA enables us to flexibly position fluorescent labels on the RNA for single-molecule fluorescence resonance energy transfer analysis and allows us to study real-time conformational dynamics during the vectorial folding process. We observed distinct signatures of the maiden secondary and tertiary folding of the Oryza sativa twister ribozyme. The maiden vectorial tertiary folding transitions occurred faster than Mg2+-induced refolding, but were also more prone to misfolding, likely due to sequential formation of alternative secondary structures. This novel assay can be applied to studying other kinetically controlled processes, such as riboswitch control and RNA-protein assembly.

SUBMITTER: Hua B

PROVIDER: S-EPMC6169520 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Publications

Mimicking Co-Transcriptional RNA Folding Using a Superhelicase.

Hua Boyang B Panja Subrata S Wang Yanbo Y Woodson Sarah A SA Ha Taekjip T

Journal of the American Chemical Society 20180803 32

Vectorial folding of RNA during transcription can produce intermediates with distinct biochemical activities. Here, we design an artificial minimal system to mimic cotranscriptional RNA folding in vitro. In this system, a presynthesized RNA molecule begins to fold from its 5'-end, as it is released from a heteroduplex by an engineered helicase that translocates on the complementary DNA strand in the 3'-to-5' direction. This chemically stabilized "superhelicase" Rep-X processively unwinds thousan ...[more]

PMID: 30063835

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Project description:We introduce and study the computational power of Oritatami, a theoretical model that explores greedy molecular folding, whereby a molecular strand begins to fold before its production is complete. This model is inspired by our recent experimental work demonstrating the construction of shapes at the nanoscale from RNA, where strands of RNA fold into programmable shapes during their transcription from an engineered sequence of synthetic DNA. In the model of Oritatami, we explore the process of folding a single-strand bit by bit in such a way that the final fold emerges as a space-time diagram of computation. One major requirement in order to compute within this model is the ability to program a single sequence to fold into different shapes dependent on the state of the surrounding inputs. Another challenge is to embed all of the computing components within a contiguous strand, and in such a way that different fold patterns of the same strand perform different functions of computation. Here, we introduce general design techniques to solve these challenges in the Oritatami model. Our main result in this direction is the demonstration of a periodic Oritatami system that folds upon itself algorithmically into a prescribed set of shapes, depending on its current local environment, and whose final folding displays the sequence of binary integers from 0 to N = 2 k - 1 with a seed of size O ( k ) . We prove that designing Oritatami is NP-hard in the number of possible local environments for the folding. Nevertheless, we provide an efficient algorithm, linear in the length of the sequence, that solves the Oritatami design problem when the number of local environments is a small fixed constant. This shows that this problem is in fact fixed parameter tractable (FPT) and can thus be solved in practice efficiently. We hope that the numerous structural strategies employed in Oritatami enabling computation will inspire new architectures for computing in RNA that take advantage of the rapid kinetic-folding of RNA.

Dataset Information

Mimicking Co-Transcriptional RNA Folding Using a Superhelicase.

Publications

Mimicking Co-Transcriptional RNA Folding Using a Superhelicase.

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