Expression, purification and characterization of the sulfite reductase hemo-subunit, SiR-HP, from Acidithiobacillus ferrooxidans

Biotechnol Lett. 2008 Jul;30(7):1239-44. doi: 10.1007/s10529-008-9679-4. Epub 2008 Mar 4.

Abstract

Sulfite reductase (SiR) is a large and soluble enzyme which catalyzes the transfer of six electrons from NADPH to sulfite to produce sulfide. The sulfite reductase flavoprotein (SiR-FP) contains both FAD and FMN, and the sulfite reductase hemoprotein (SiR-HP) contains an iron-sulfur cluster coupled to a siroheme. The enzyme is arranged so that the redox cofactors in the FAD-FMN-Fe(4)S(4)-Heme sequence make an electron pathway between NADPH and sulfite. Here we report the cloning, expression, and characterization of the SiR-HP of the sulfite reductase from Acidithiobacillus ferrooxidans. The purified SiR-HP contained a [Fe(4)S(4)] cluster. Site-directed mutagenesis results revealed that Cys427, Cys433, Cys472 and Cys476 were in ligating with the [Fe(4)S(4)] cluster of the protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acidithiobacillus / enzymology*
  • Acidithiobacillus / genetics*
  • Amino Acid Substitution
  • Bacterial Proteins / biosynthesis*
  • Bacterial Proteins / genetics*
  • Cloning, Molecular / methods
  • Cysteine / genetics
  • Cysteine / metabolism
  • Mutagenesis, Site-Directed / methods
  • Mutation, Missense
  • Sulfite Reductase (NADPH) / biosynthesis*
  • Sulfite Reductase (NADPH) / genetics*

Substances

  • Bacterial Proteins
  • Sulfite Reductase (NADPH)
  • Cysteine