Expression, purification and characterization of the sulfite reductase hemo-subunit, SiR-HP, from Acidithiobacillus ferrooxidans.
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.
Zeng
Jia
J
Department of Bioengineering, School of Resources Processing and Bioengineering, Central South University, Changsha, 410083, People's Republic of China. csuzengjia@yahoo.com.cn
Wang
Ming
M
Zhang
Xiaojian
X
Wang
Yiping
Y
Ai
Chenbin
C
Liu
Jianshe
J
Qiu
Guanzhou
G
D042763:Acidithiobacillus / Q000201:enzymology* / Q000235:genetics*; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D016297:Mutagenesis, Site-Directed / Q000379:methods; D020125:Mutation, Missense; D050864:Sulfite Reductase (NADPH) / Q000096:biosynthesis* / Q000235:genetics*
D001426:Bacterial Proteins; D050864:Sulfite Reductase (NADPH); D003545:Cysteine