XpsE oligomerization triggered by ATP binding, not hydrolysis, leads to its association with XpsL

Abstract

GspE belongs to a secretion NTPase superfamily, members of which are involved in type II/IV secretion, type IV pilus biogenesis and DNA transport in conjugation or natural transformation. Predicted to be a cytoplasmic protein, GspE has nonetheless been shown to be membrane‐associated by interacting with the N‐terminal cytoplasmic domain of GspL. By taking biochemical and genetic approaches, we observed that ATP binding triggers oligomerization of Xanthomonas campestris XpsE (a GspE homolog) as well as its association with the N‐terminal domain of XpsL (a GspL homolog). While isolated XpsE exhibits very low intrinsic ATPase activity, association with XpsL appears to stimulate ATP hydrolysis. Mutation at a conserved lysine residue in the XpsE Walker A motif causes reduction in its ATPase activity without significantly influencing its interaction with XpsL, congruent with the notion that XpsE–XpsL association precedes ATP hydrolysis. For the first time, functional significance of ATP binding to GspE in type II secretion system is clearly demonstrated. The implications may also be applicable to type IV pilus biogenesis.

Zinc and ATP Binding of the Hexameric AAA-ATPase PilF from Thermus thermophilus

“…The assembly of some well investigated polymerization ATPases, such as GspE from Vibrio cholerae and TrbB from Helicobacter pylori requires ATP (34,35). In contrast the assembly of the PilF complex in T. thermophilus does not require ATP.…”

Interaction domains in the Pseudomonas aeruginosa type II secretory apparatus component XcpS (GspF)

“…XcpR contains a conserved Walker A-box motif that was shown to be indispensable for its function (Turner et al, 1993). Binding of ATP was recently shown to trigger oligomerization of the X. campestris XcpR homologue XpsE (Shiue et al, 2006) probably into hexamers (Crowther et al, 2005;Savvides et al, 2003). Knowledge of the role of XcpS in the secreton and its interactions with other Xcp components is rather limited.…”

Mutation of a key residue in the type II secretion system ATPase uncouples ATP hydrolysis from protein translocation

“…Co-crystallization of the N-terminal region of EpsE and the cytoplasmic domain of EpsL (Abendroth et al, 2005) further supports the proposition that the N-terminal region of GspE interacts directly with the cytoplasmic domain of GspL. Shiue et al (2006) recently demonstrated that the X. campestris pv. campestris XpsE associated with the cytoplasmic domain of XpsL as hexamer in an ATP binding-dependent manner.…”

“…Not until recently, ATPase activity, albeit weak, was demonstrated in both the V. cholerae EpsE and the X. campestris pv. campestris XpsE (Camberg and Sandkvist, 2005;Shiue et al, 2006). Each was stimulated by cytoplasmic domain of its respective interactive partner EpsL or XpsL (Shiue et al, 2006;Camberg et al, 2007).…”