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A thiamin-utilizing ribozyme decarboxylates a pyruvate-like substrate

Nature Chemistry volume 5, pages 971–977 (2013)Cite this article

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Abstract

Vitamins are hypothesized to be relics of an RNA world, and were probably participants in RNA-mediated primordial metabolism. If catalytic RNAs, or ribozymes, could harness vitamin cofactors to aid their function in a manner similar to protein enzymes, it would enable them to catalyse a much larger set of chemical reactions. The cofactor thiamin diphosphate, a derivative of vitamin B1 (thiamin), is used by enzymes to catalyse difficult metabolic reactions, including decarboxylation of stable α-keto acids such as pyruvate. Here, we report a ribozyme that uses free thiamin to decarboxylate a pyruvate-based suicide substrate (LnkPB). Thiamin conjugated to biotin was used to isolate catalytic individuals from a pool of random-sequence RNAs attached to LnkPB. Analysis of a stable guanosine adduct obtained via digestion of an RNA sequence (clone dc4) showed the expected decarboxylation product. The discovery of a prototypic thiamin-utilizing ribozyme has implications for the role of RNA in orchestrating early metabolic cycles.

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Figure 1: A strategy for isolating ribozymes with α-keto acid decarboxylase activity.
Figure 2: In vitro selection of decarboxylase ribozymes.
Figure 3: Mass spectroscopy of the decarboxylation product.
Figure 4: Unmodified thiamin is an effective cofactor

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Acknowledgements

This work was supported by the Natural Sciences and Engineering Research Council of Canada (NSERC). The authors thank R. N. Young and the members of his group for advice and assistance with organic synthesis, and N. S. Kumar, A. J. Bennet, P. J. Unrau, E. C. Young, A. R. Lewis and H. Chen for their expert and practical advice.

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Authors and Affiliations

  1. Department of Molecular Biology & Biochemistry, Simon Fraser University, Burnaby, V5A 1S6, British Columbia, Canada

    Paul Cernak & Dipankar Sen

  2. Department of Chemistry, Simon Fraser University, Burnaby, V5A 1S6, British Columbia, Canada

    Dipankar Sen

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  1. Paul Cernak
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Contributions

P.C. and D.S. conceived and designed the experiments. P.C. performed the experiments. P.C. and D.S. analysed the data. P.C. and D.S. co-wrote the paper.

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Correspondence to Dipankar Sen.

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Cernak, P., Sen, D. A thiamin-utilizing ribozyme decarboxylates a pyruvate-like substrate. Nature Chem 5, 971–977 (2013). https://doi.org/10.1038/nchem.1777

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