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Discovery of proteinaceous N-modification in lysine biosynthesis of Thermus thermophilus

Nature Chemical Biology volume 5, pages 673–679 (2009)Cite this article

Abstract

Although the latter portion of lysine biosynthesis, the conversion of α-aminoadipate (AAA) to lysine, in Thermus thermophilus is similar to the latter portion of arginine biosynthesis, enzymes homologous to ArgA and ArgJ are absent from the lysine pathway. Because ArgA and ArgJ are known to modify the amino group of glutamate to avoid intramolecular cyclization of intermediates, their absence suggests that the pathway includes an alternative N-modification system. We reconstituted the conversion of AAA to lysine and found that the amino group of AAA is modified by attachment to the γ-carboxyl group of the C-terminal Glu54 of a small protein, LysW; that the side chain of AAA is converted to the lysyl side chain while still attached to LysW; and that lysine is subsequently liberated from the LysW-lysine fusion. The fact that biosynthetic enzymes recognize the acidic globular domain of LysW indicates that LysW acts as a carrier protein or protein scaffold for the biosynthetic enzymes. This study thus reveals the previously unknown function of a small protein in primary metabolism.

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Figure 1: Lysine biosynthetic pathway.
Figure 2: Gene clusters for lysine biosynthetic enzymes and carrier proteins in T. thermophilus, D. radiodurans, and several archaea.
Figure 3: In vitro lysine synthesis.
Figure 4: Analysis of synthetic intermediates.
Figure 5: γ-linkage-specific reactions of lysine biosynthetic enzymes.
Figure 6: Proposed LysW-mediated mechanism of lysine biosynthesis.

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Acknowledgements

We thank H. Satsu for technical help with amino acid analysis. This work was supported in part by a grant-in-aid for scientific research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan, from the Nagase Science and Technology Foundation, from the Asahi Glass Foundation and from the Charitable Trust Araki Medical and Biochemistry Memorial Research Promotion Fund.

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Author notes

  1. Akira Horie & Asako Saiki

    Present address: Present addresses: Central Laboratories for Frontier Technology, Kirin Holdings Co., Yokohama, Japan (A.H); Life Science Division, Yamaha Motor Co., Shizuoka, Japan (A.S.).,

Authors and Affiliations

  1. Biotechnology Research Center, The University of Tokyo, Bunkyo-ku, Tokyo, Japan

    Akira Horie, Takeo Tomita, Asako Saiki, Tomohisa Kuzuyama & Makoto Nishiyama

  2. Computational Biology Group, Quantum Beam Science Directorate, Japan Atomic Energy Agency, Kizugawa, Kyoto, Japan

    Hidetoshi Kono

  3. Quantum Bioinformatics Team, Center for Computational Science and e-System, Japan Atomic Energy Agency, Kizugawa, Kyoto, Japan

    Hidetoshi Kono

  4. Division of Biochemical Analysis, Central Laboratory of Medical Sciences, Juntendo University School of Medicine, Bunkyo-ku, Tokyo, Japan

    Hikari Taka, Reiko Mineki & Tsutomu Fujimura

  5. Atopy (Allergy) Research Center, Juntendo University School of Medicine, Bunkyo-ku, Tokyo, Japan

    Chiharu Nishiyama

  6. RIKEN SPring-8 Center, Sayo-cho, Sayo-gun, Hyogo, Japan

    Makoto Nishiyama

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Contributions

Research planning and supervision were by T.T., T.K. and M.N.; biochemical experiments were by A.H.; gene knockout and replacement of T. thermophilus were by A.S.; LC-MS/MS and MALDI-TOF MS were by H.T., R.M., T.F. and C.N.; in silico modeling was by H.K. and T.T.; and manuscript writing was by A.H., T.T. and M.N.

Corresponding author

Correspondence to Makoto Nishiyama.

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Supplementary Figures 1–9, Supplementary Tables 1 and 2 and Supplementary Methods (PDF 4638 kb)

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Horie, A., Tomita, T., Saiki, A. et al. Discovery of proteinaceous N-modification in lysine biosynthesis of Thermus thermophilus. Nat Chem Biol 5, 673–679 (2009). https://doi.org/10.1038/nchembio.198

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