Nucleotide-dependent conformational changes in a protease-associated ATPase HsIU

Abstract

Background: 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 HsIV peptidase and forms an ATP-dependent HsIVU protease. Results: We have characterized four distinct HsIU conformational states, going 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 nucleotide identity in one structure and reveals the ATP hydrolysis-dependent structural changes in the HsIVU 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.