Crystal Structure of a Type IV Pilus Assembly ATPase: Insights into the Molecular Mechanism of PilB from Thermus thermophilus

Type IV pili (T4P) mediate bacterial motility and virulence. The PilB/GspE family ATPases power the assembly of T4P and type 2 secretion systems. We determined the structure of the ATPase region of PilB (PilBATP) in complex with ATPγS to provide a model of a T4P assembly ATPase and a view of a PilB/GspE family hexamer at better than 3-Å resolution. Spatial positioning and conformations of the protomers suggest a mechanism of force generation. All six PilBATP protomers contain bound ATPγS. Two protomers form a closed conformation poised for ATP hydrolysis. The other four molecules assume an open conformation but separate into two pairs with distinct active-site accessibilities. We propose that one pair represents the post-hydrolysis phase while the other pair appears poised for ADP/ATP exchange. Collectively, the data suggest that T4P assembly is powered by coordinating concurrent substrate binding with ATP hydrolysis across the PilB hexamer. •Crystal structure of a type IV pilus assembly ATPase•PilB in complex with ATPγS assumes three distinct positions within the hexamer•Binding studies suggest that ADP is actively displaced by ATP during catalysis•Ring positions indicate a symmetric rotary catalytic mechanism of force generation Hexameric ATPases power numerous cellular activities critical for bacterial virulence. Mancl et al. have solved the structure of a type IV assembly ATPase to produce insights into the mechanism of force generation and broadly into the mechanism of catalysis of PilB/GspE family enzymes.