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A cyclic form of N6-threonylcarbamoyladenosine as a widely distributed tRNA hypermodification

Nature Chemical Biology volume 9, pages 105–111 (2013)Cite this article

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Abstract

N6-threonylcarbamoyladenosine (t6A) is a universally conserved, essential modified nucleoside found in transfer RNAs (tRNAs) responsible for ANN codons in all three domains of life. t6A has a crucial role in maintaining decoding accuracy during protein synthesis. The presence of t6A in cellular tRNAs has been well documented for more than four decades. However, under conditions optimized for nucleoside preparation, we detected little t6A in tRNAs from Escherichia coli. Instead, we identified a new modified base named 'cyclic t6A' (ct6A), which is a cyclized active ester with an oxazolone ring. An E1-like enzyme, CsdL (renamed as TcdA), which catalyzes ATP-dependent dehydration of t6A to form ct6A, was also identified. Two yeast homologs of tcdA, YHR003C (TCD1) and YKL027W (TCD2), were required for ct6A formation and respiratory cell growth. ct6A was involved in promoting decoding efficiency. Structural modeling suggests that ct6A recognizes the first adenine base of ANN codon at the ribosomal A site.

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Figure 1: Identification of a new modified nucleoside N394 at position 37 of E. coli tRNA.
Figure 2: Structural determination of N394.
Figure 3: Identification of TcdA and in vitro reconstitution of ct6A.
Figure 4: Yeast homologs responsible for ct6A formation are required for respiratory cell growth.
Figure 5: Decoding property of ct6A.

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Acknowledgements

We are grateful to the members of the Suzuki laboratory, especially T. Suzuki, A. Nagao, Y. Sakaguchi, T. Ohira, M. Ohara, T. Chujo, H. Takeda, Y. Ikeuchi, T. Taniguchi and T. Sakashita for technical assistance and fruitful discussions. Special thanks are due to R.W. Adamiak (Polish Academy of Sciences) for providing t6A. This work was supported by the Grants-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Science, Sports and Culture of Japan (to T.S.) and by a grant from the New Energy and Industrial Technology Development Organization (to T.S.).

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  1. Department of Chemistry and Biotechnology, Graduate School of Engineering, the University of Tokyo, Tokyo, Japan

    Kenjyo Miyauchi, Satoshi Kimura & Tsutomu Suzuki

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  1. Kenjyo Miyauchi
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  2. Satoshi Kimura
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Contributions

K.M. and T.S. designed this study. K.M. and S.K. performed the series of experiments. K.M. and T.S. wrote this paper. T.S. supervised all work.

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Correspondence to Tsutomu Suzuki.

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Supplementary Data Set 1

Candidate genes for ct6A formation selected by the Integrated Microbial Genomes (IMG) system (XLS 0 kb)

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Miyauchi, K., Kimura, S. & Suzuki, T. A cyclic form of N6-threonylcarbamoyladenosine as a widely distributed tRNA hypermodification. Nat Chem Biol 9, 105–111 (2013). https://doi.org/10.1038/nchembio.1137

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