A model system for [NiFe] hydrogenase maturation studies: Purification of an active site-containing hydrogenase large subunit without small subunit

Gordon Winter, Thorsten Buhrke, Oliver Lenz, Anne Katherine Jones, Michael Forgber, Bärbel Friedrich

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The large subunit HoxC of the H2-sensing [NiFe] hydrogenase from Ralstonia eutropha was purified without its small subunit. Two forms of HoxC were identified. Both forms contained iron but only substoichiometric amounts of nickel. One form was a homodimer of HoxC whereas the second also contained the Ni-Fe site maturation proteins HypC and HypB. Despite the presence of the Ni-Fe active site in some of the proteins, both forms, which lack the Fe-S clusters normally present in hydrogenases, cannot activate hydrogen. The incomplete insertion of nickel into the Ni-Fe site provides direct evidence that Fe precedes Ni in the course of metal center assembly.

Original languageEnglish
Pages (from-to)4292-4296
Number of pages5
JournalFEBS Letters
Issue number20
Publication statusPublished - Aug 15 2005



  • Maturation
  • Metal center assembly
  • Ralstonia eutropha
  • [NiFe] hydrogenase

ASJC Scopus subject areas

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

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