Article,
Evidence that the Streptomyces developmental protein WhiD, a member of the WhiB family, binds a 4Fe-4S cluster.
J Biol Chem, 280 (9): 8309--8315 (March 2005)
DOI: 10.1074/jbc.M412622200
Abstract
WhiD is required for the late stages of sporulation in the Gram-positive
bacterium Streptomyces coelicolor. WhiD is a member of the WhiB-like
family of putative transcription factors that are present throughout
the actinomycetes but absent from other organisms. This family of
proteins has four near-invariant cysteines, suggesting that these
residues might act as ligands for a metal cofactor. Overexpressed
WhiD, purified from Escherichia coli, contained substoichiometric
amounts of iron and had an absorption spectrum characteristic of
a 2Fe-2S cluster. After Fe-S cluster reconstitution under anaerobic
conditions, WhiD contained approximately 4 iron atoms/monomer and
similar amounts of sulfide ion and gave an absorption spectrum characteristic
of a 4Fe-4S cluster. Reconstituted WhiD gave no electron paramagnetic
resonance signal as prepared but, after reduction with dithionite,
gave an electron paramagnetic resonance signal (g approximately
2.06, 1.94) consistent with a one-electron reduction of a 4Fe-4S(2+)
cluster to a 4Fe-4S(1+) state with electron spin of S = (1/2).
The anaerobically reconstituted 4Fe-4S cluster was oxygen sensitive.
Upon exposure to air, absorption at 410 and 505 nm first increased
and then showed a steady decrease with time until the protein was
colorless in the near UV/visible region. These changes are consistent
with an oxygen-induced change from a 4Fe-4S to a 2Fe-2S cluster,
followed by complete loss of cluster from the protein. Each of the
four conserved cysteine residues, Cys-23, -53, -56, and -62, was
essential for WhiD function in vivo.
