20.109(F09):Stephanie Bachar and Matt Luchette

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1. Background Information

  • Synthetic biological systems can be used to perform a predictable task
  • Recently, scientists have designed modified amino acids for use in protein synthesis
  • In order to incorporate these amino acids, tyrosyl-tRNA synthase must also be modified.
  • Similar projects have developed unnatural amino acids and modified tyrosyl-tRNAs that translate at varying efficiencies. Examples include:

Fuwa, T. & Miyazawa, T. (1988) Proc. Natl. Acad. Sci. USA 85, 6237–6241.

    • Hendrickson, W. A., Horton, J. R. & LeMaster, D. M. (1990) EMBO J. 9,

1665–1672.

    • Yabuki, T., Kigawa, T., Dohmae, N., Takio, K., Terada, T., Ito, Y., Laue, E. D.,

Cooper, J. A., Kainosho, M. & Yokoyama, S. (1998) J. Biomol. NMR 11, 295–306.

    • Hirao, I., Ohtsuki, T., Fujiwara, T., Mitsui, T., Yokogawa, T., Okuni, T., Nakayama,

H., Takio, K., Yabuki, T., Kigawa, T., et al. (2002) Nat. Biotechnol. 20, 177–182.

    • Koide, H., Yokoyama, S., Katayama, Y., Muto, Y., Kigawa, T., Kohno, T.,

Takusari, H., Oishi, M., Takahashi, S., Tsukumo, K., et al. (1994) Biochemistry 33, 7470–7476.

    • Kiick, K. L., Weberskirch, R. & Tirrell, D. A. (2001) FEBS Lett. 502, 25–30.
    • Kigawa, T., Yamaguchi-Nunokawa, E., Kodama, K., Matsuda, T., Yabuki, T.,

Matusda, N., Ishitani, R., Nureki, O. & Yokoyama, S. (2001) J. Struct. Funct. Genomics 2, 27–33.

2. Our Idea

  • These papers have shown tRNA synthases that incorporated unnatural amino acids into a protein at varying efficiencies. nthWe think that by either examining the tRNA synthases or by preforming our own syntheses and screenings of unique tRNA synthases, we can better examine residues in these proteins that are key for specificity of our mutated tRNA synthases.

To do this, we must first gather existing knowledge on how natural tRNA synthases bind to and incorporate natural amino acids into a protein. Have these key amino acid binding residues already been identified? Furthermore,

3. Applications