Signal recognition particle-dependent inner membrane targeting of the PulG Pseudopilin component of a type II secretion system.

Olivera Francetic (1) , Nienke Buddelmeijer (1) , Shawn Lewenza (1) , Carol A Kumamoto (2) , Anthony P Pugsley (1)

1 Génétique Moléculaire
2 Tufts University School of Medicine [Boston]

The pseudopilin PulG is an essential component of the pullulanase-specific type II secretion system from Klebsiella oxytoca. PulG is the major subunit of a short, thin-filament pseudopilus, which presumably elongates and retracts in the periplasm, acting as a dynamic piston to promote pullulanase secretion. It has a signal sequence-like N-terminal segment that, according to studies with green and red fluorescent protein chimeras, anchors unassembled PulG in the inner membrane. We analyzed the early steps of PulG inner membrane targeting and insertion in Escherichia coli derivatives defective in different protein targeting and export factors. The beta-galactosidase activity in strains producing a PulG-LacZ hybrid protein increased substantially when the dsbA, dsbB, or all sec genes tested except secB were compromised by mutations. To facilitate analysis of native PulG membrane insertion, a leader peptidase cleavage site was engineered downstream from the N-terminal transmembrane segment (PrePulG*). Unprocessed PrePulG* was detected in strains carrying mutations in secA, secY, secE, and secD genes, including some novel alleles of secY and secD. Furthermore, depletion of the Ffh component of the signal recognition particle (SRP) completely abolished PrePulG* processing, without affecting the Sec-dependent export of periplasmic MalE and RbsB proteins. Thus, PulG is cotranslationally targeted to the inner membrane Sec translocase by SRP.

Domaines: Bactériologie

Olivera Francetic, Nienke Buddelmeijer, Shawn Lewenza, Carol A Kumamoto, Anthony P Pugsley. Signal recognition particle-dependent inner membrane targeting of the PulG Pseudopilin component of a type II secretion system.. Journal of Bacteriology, 2007, 189 (5), pp.1783-1793. ⟨10.1128/JB.01230-06⟩. ⟨hal-00138935⟩