Nucleotide-Dependent Conformational Changes in a Protease-Associated ATPase HslU

AbstractBackground: The bacterial heat shock locus ATPase HslU is an AAA+ protein that has structures known in many nucleotide-free and -bound states. Nucleotide is required for the formation of the biologically active HslU hexameric assembly. The hexameric HslU ATPase binds the dodecameric HslV peptidase and forms an ATP-dependent HslVU protease.Results: We have characterized four distinct HslU conformational states, going sequentially from open to closed: the empty, SO4, ATP, and ADP states. The nucleotide binds at a cleft formed by an α/β domain and an α-helical domain in HslU. The four HslU states differ by a rotation of the α-helical domain. This classification leads to a correction of nucleotide identity in one structure and reveals the ATP hydrolysis-dependent structural changes in the HslVU complex, including a ring rotation and a conformational change of the HslU C terminus. This leads to an amended protein unfolding-coupled translocation mechanism.Conclusions: The observed nucleotide-dependent conformational changes in HslU and their governing principles provide a framework for the mechanistic understanding of other AAA+ proteins