Characterization of Mycobacterium tuberculosis WhiB1/Rv3219 as a protein disulfide reductase.
WhiB family of protein is emerging as one of the most fascinating group and is implicated in stress response as well as pathogenesis via their involvement in diverse cellular processes. Surprisingly, available in vivo data indicate an organism specific physiological role for each of these proteins. The WhiB proteins have four conserved cysteine residues where two of them are present in a C-X-X-C motif. In thioredoxins and similar proteins, this motif works as an active site and confers thiol-disulfide oxidoreductase activity to the protein. The recombinant WhiB1/Rv3219 was purified in a single step from Escherichia coli using Ni(2+)-NTA affinity chromatography and was found to exist as a homodimer. Mass spectrometry of WhiB1 shows that the four cysteine residues form two intramolecular disulfide bonds. Using intrinsic tryptophan fluorescence as a measure of redox state, the redox potential of WhiB1 was calculated as -236+/-2mV, which corresponds to the redox potential of many cytoplasmic thioredoxin-like proteins. WhiB1 catalyzed the reduction of insulin disulfide thus clearly demonstrating that it functions as a protein disulfide reductase. Present study for the first time suggests that WhiB1 may be a part of the redox network of Mycobacterium tuberculosis through its involvement in thiol-disulfide exchange with other cellular proteins.
Garg
Saurabh K
SK
Institute of Microbial Technology, Sector 39A, Chandigarh 160 036, India.
Suhail Alam
Md
M
Soni
Vishal
V
Radha Kishan
K V
KV
Agrawal
Pushpa
P
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D003545:Cysteine / Q000737:chemistry; D015870:Gene Expression; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000737:chemistry / Q000201:enzymology*; D011490:Protein Disulfide Reductase (Glutathione) / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013879:Thioredoxins / Q000737:chemistry
D013879:Thioredoxins; D011490:Protein Disulfide Reductase (Glutathione); D003545:Cysteine