Interaction of the iron-sulfur cluster assembly protein IscU with the Hsc66/Hsc20 molecular chaperone system of Escherichia coli

The iscU gene in bacteria is located in a gene cluster encoding proteins im plicated in iron-sulfur cluster assembly and an hsc70-type (heat shock cogn ate) molecular chaperone system, iscSUA-hscBA. To investigate possible inte ractions between these systems, we have overproduced and purified the IscU protein from Escherichia coli and have studied its interactions with the hs cA and hscB gene products Hsc66 and Hsc20. IscU and its iron-sulfur complex (IscU-Fe/S) stimulated the basal steady-state ATPase activity of Hsc66 wea kly in the absence of Hsc20 but, in the presence of Hsc20, increased the AT Pase activity up to 480-fold. Hsc20 also decreased the apparent K-m for Isc U stimulation of Hsc66 ATPase activity, and surface plasmon resonance studi es revealed that Hsc20 enhances binding of IscU to Hsc66. Surface plasmon r esonance and isothermal titration calorimetry further showed that IscU and Hsc20 form a complex, and Hsc20 may thereby aid in the targeting of IscU to Hsc66. These results establish a direct and specific role for the Hsc66/Hs c20 chaperone system in functioning with isc gene components for the assemb ly of iron-sulfur cluster proteins.