Interaction between the autokinase EpsE and EpsL in the cytoplasmic membrane is required for extracellular secretion in Vibrio cholerae.

Abstract

Vibrio cholerae secretes a number of proteins important for virulence, including cholera toxin. This process requires the products of the eps genes which have homologues in genera such as Aeromonas, Klebsiella and Pseudomonas and are thought to form a membrane‐associated multiprotein complex. Here we show that the putative nucleotide‐binding protein EpsE is associated with and stabilized by the cytoplasmic membrane via interaction with EpsL. Analysis of fusion proteins between EpsE and the homologous ExeE from Aeromonas hydrophila demonstrates that the N‐terminus of EpsE contains the EpsL binding domain and determines species specificity. An intact Walker A box, commonly found in ATP‐binding proteins, is required for activity of EpsE in vivo and for autophosphorylation of purified EpsE in vitro. These results indicate that both the kinase activity of EpsE as well as its ability to interact with the putative cytoplasmic membrane protein EpsL are required for translocation of toxin across the outer membrane in Vibrio cholerae.

A membrane metalloprotease participates in the sequential degradation of a Caulobacter polarity determinant

“…Separation of C. crescentus cell lysates into soluble and membrane fractions was performed as previously described (Sandkvist et al, 1995;Viollier et al, 2002a). Cells were grown in rich media until OD 600 reached 0.7-0.8.…”

Cell surface localization and processing of the ComG proteins, required for DNA binding during transformation of Bacillus subtilis

“…1A), demonstrating that this protein behaves as a peripheral membrane protein (Russel and Model, 1982). Indeed, hydropathy analysis of ComGA had not predicted the presence of a membrane-spanning segment (Albano et al, 1989), and at least one other member of the ComGA family has also been shown to be a peripheral membrane protein (Sandkvist et al, 1995).…”

Overproduction of the secretin OutD suppresses the secretion defect of an Erwinia chrysanthemi outB mutant

“…Their leader peptide is cleaved by the GspO prepilin peptidase (Nunn & G. CONDEMINE and V. E. SHEVCHIK 1991). GspE is an ATP-binding protein with a kinase activity (Sandkvist et al, 1995). The bitopic innermembrane proteins GspC, L and M have a large periplasmic domain and GspF is a polytopic innermembrane protein (Reeves et al, 1994 ;Thomas et al, 1997).…”

“…The bitopic innermembrane proteins GspC, L and M have a large periplasmic domain and GspF is a polytopic innermembrane protein (Reeves et al, 1994 ;Thomas et al, 1997). Some possible interactions between components of the GSP machinery have been described : OutE-OutL, EspE-EspL, XcpT-XcpR, XcpY-XcpZ and EpsLEpsM (Py et al, 1999 ;Sandkvist et al, 1995Sandkvist et al, , 1999Kagami et al, 1998 ;Michel et al, 1998).…”