Background: The bacterial heat shock locus ATPase HslU is an AAA(+) protein
that has structures known in many nucleotide-free and -bound states. Nucle
otide is required for the formation of the biologically active HslU hexamer
ic assembly. The hexameric HslU ATPase binds the dodecameric HsIV peptidase
and forms an ATP-dependent HsIVU protease.
Results: We have characterized four distinct HsIU conformational states, go
ing sequentially from open to closed: the empty, SO4, ATP, and ADP states.
The nucleotide binds at a cleft formed by an alpha/beta domain and an alpha
-helical domain in HslU. The four HslU states differ by a rotation of the
alpha -helical domain. This classification leads to a correction of nucleot
ide identity in one structure and reveals the ATP hydrolysis-dependent stru
ctural changes in the HsIVU complex, including a ring rotation and a confor
mational change of the HslU C terminus. This leads to an amended protein un
folding-coupled translocation mechanism.
Conclusions: The observed nucleotide-dependent conformational changes in Hs
lU and their governing principles provide a framework for the mechanistic u
nderstanding of other AAA(+) proteins.