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

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.