IscU is a scaffold
protein that functions in
iron-
sulfur cluster assembly and transfer. Its critical importance has been recently underscored by the finding that a single intronic mutation in the human
iscu gene is associated with a
myopathy resulting from deficient
succinate dehydrogenase and
aconitase [Mochel, F., Knight, M. A., Tong, W. H., Hernandez, D., Ayyad, K., Taivassalo, T., Andersen, P. M., Singleton, A., Rouault, T. A., Fischbeck, K. H., and Haller, R. G. (2008) Am. J. Hum. Genet. 82, 652-660].
IscU functions through interactions with a chaperone
protein HscA and a cochaperone
protein HscB. To probe the molecular basis for these interactions, we have used NMR spectroscopy to investigate the
solution structure of
IscU from Escherichia coli and its interaction with HscB from the same organism. We found that wild-type apo-
IscU in
solution exists as two distinct conformations: one largely disordered and one largely ordered except for the
metal binding residues. The two states interconvert on the millisecond time scale. The ordered conformation is stabilized by the addition of
zinc or by the single-site
IscU mutation, D39A. We used apo-IscU(D39A) as a surrogate for the folded state of wild-type
IscU and assigned its NMR spectrum. These assignments made it possible to identify the region of
IscU with the largest structural differences in the two conformational states. Subsequently, by following the NMR signals of apo-IscU(D39A) upon addition of HscB, we identified the most perturbed regions as the two N-terminal beta-strands and the C-terminal alpha-helix. On the basis of these results and analysis of
IscU sequences from multiple species, we have identified the surface region of
IscU that interacts with HscB. We conclude that the
IscU-HscB complex exists as two (or more) distinct states that interconvert at a rate much faster than the rate of dissociation of the complex and that HscB binds to and stabilizes the ordered state of apo-
IscU.