Functional expression of thiocyanate hydrolase is promoted by its activator protein, P15K

Shingo Kataoka, Takatoshi Arakawa, Shota Hori, Yoko Katayama, Yoshiko Hara, Yasuhiko Matsushita, Hiroshi Nakayama, Masafumi Yohda, Hiroshi Nyunoya, Naoshi Dohmae, Mizuo Maeda, Masafumi Odaka

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Abstract

Thiocyanate hydrolase (SCNase) is a cobalt-containing enzyme with a post-translationally modified cysteine ligand, γCys131-SO2H. When the SCNase α, β and γ subunits were expressed in Escherichia coli, the subunits assembled to form a hetero-dodecamer, (αβγ)4, like native SCNase but exhibited no catalytic activity. Metal analysis indicated that SCNase was expressed as an apo-form irrespective of the presence of cobalt in the medium. On the contrary, SCNase co-expressed with P15K, encoded just downstream of SCNase genes, in cobalt-enriched medium under the optimized condition (SCNase(+P15K)) possessed 0.86 Co atom/αβγ trimer and exhibited 78% of the activity of native SCNase. SCNase(+P15K) showed a UV-Vis absorption peak characteristic of the SCNase cobalt center. About 70% of SCNase(+P15K) had the γCys131-SO2H modification. These results indicate that SCNase(+P15K) is the active holo-SCNase. P15K is likely to promote the functional expression of SCNase probably by assisting the incorporation of cobalt ion.

Original languageEnglish
Pages (from-to)4667-4672
Number of pages6
JournalFEBS Letters
Volume580
Issue number19
DOIs
Publication statusPublished - 2006 Aug 21

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Keywords

  • Activator protein
  • Apo-protein
  • Cysteine-sulfenic acid
  • Cysteine-sulfinic acid
  • Nitrile hydratase
  • Non-corrin cobalt
  • Post-translational modification
  • Thiocyanate hydrolase

ASJC Scopus subject areas

  • Biophysics
  • Structural Biology
  • Biochemistry
  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

Kataoka, S., Arakawa, T., Hori, S., Katayama, Y., Hara, Y., Matsushita, Y., Nakayama, H., Yohda, M., Nyunoya, H., Dohmae, N., Maeda, M., & Odaka, M. (2006). Functional expression of thiocyanate hydrolase is promoted by its activator protein, P15K. FEBS Letters, 580(19), 4667-4672. https://doi.org/10.1016/j.febslet.2006.07.051