1.B.17 The Outer Membrane Factor (OMF) Family

Proteins of the OMF family (Li et al., 2001; Wong et al., 2001) function in conjunction with a primary cytoplasmic membrane transporter of the MFS (TC #2.A.1) (Pao et al., 1998; Wang et al. 2020), the ABC superfamily (TC #3.A.1) (Saurin et al., 1999), the RND superfamily (TC #2.A.6) (Tseng et al., 1999) and the ArAT family (TC #2.A.85) (Harley and Saier, 2000) as well as a membrane fusion protein (MFP; TC #8.A.1) (Dinh et al., 1994). The complex thus formed allows transport (export) of various solutes (heavy metal cations; drugs, oligosaccharides, proteins, etc.) across the two envelopes of the Gram-negative bacterial cell envelope in a single energy-coupled step. The OMF proteins probably form homotrimeric 12 stranded β-barrel-type pores in the outer membrane through which the solutes pumped out of the cytoplasm or cytoplasmic membrane pass in response to the energy-coupled export process catalyzed by the cytoplasmic membrane permease. In one case, the complex of primary transporter, MFP and OMF forms transiently in response to substrate binding (Létoffé et al., 1996). In another case involving AcrA (RND superfamily; TC#s 2.A.6.2.2;  8.A.1.6.1) and TolC (1.B.17.1.1), the interaction appears to be substrate independent (Husain et al., 2004).

The Serratia marcescens hemophore is secreted by a type I (ABC) secretion system consisting of three proteins: a membrane ABC protein, an adaptor protein, and the TolC-like outer membrane factor (Cescau et al., 2007). Assembly of these proteins is induced by substrate binding to the ABC protein. A hemophore mutant lacking the last 14 C-terminal amino acids is not secreted but rather interacts with the ABC protein and promotes a stable multiprotein complex. Strains expressing the transporter and the mutant protein are sensitive to detergents (e.g., sodium dodecyl sulfate [SDS]). TolC is trapped in the transporter, jammed by the truncated substrate, and therefore is not present at sufficient concentrations to allow the efflux pumps to expel detergents. Using an SDS sensitivity assay, the hemophore proved to interact with the ABC protein via two non-overlapping sites. The C-terminal peptide, which functions as an intramolecular signal sequence in the complete substrate, may have intermolecular activity and trigger complex dissociation (Cescau et al., 2007).

The crystal structure of E. coli TolC has been solved to 2.1 Å resolution (Koronakis et al., 1997, 2000), and the VceC homologue of Vibrio cholerae has been solved to 1.8 Å resolution (Federici et al., 2005). Three TolC protomers form a continuous, solvent-accessible conduit, a channel tunnel over 140 Å long that spans both the outer membrane (as 12 β-strands, 4 each per protomer) and the periplasmic space (as 12 α-helices, 6 continuous, 6 discontinuous, 4 each protomer). The α-helices are continuous with the β-strands. The periplasmic end of the tunnel is sealed by sets of coiled helices that might untwist upon contact with the primary permease to open the channel (Andersen et al., 2001; Koronakis et al., 2001). 3-d structures of sequential open states in a symmetrical opening transition of the TolC of E. coli exit duct have been identified (Pei et al., 2011).

The OMFs exhibit a preudosymmetrical structure due to the presence of two internally duplicated segments. Thus, the outer membrane β-barrel is assembled from the three protomers with each one contributing 4 β-strands. Each strand is between 10 and 13 residues long. The strands both curve and twist, yielding a superhelical structure, but the channel is wide open and fully accessible to solvent. The possibility of channel closure due to conformational mobility has not been excluded (Koronakis et al., 2000). The results clearly suggest that the OMF (and not the MFP) is largely responsible for the formation of both the trans-outer membrane and trans-periplasmic channels.

OMF family members are found in most classes of proteobacteria, in cyanobacteria, spirochetes, and in species of Deinococcus, Aquafex and Porphyromonas. The proteins are of 347-541 aas in length and are exported to the outer membrane via the general secretory pathway (GSP; TC#3.A.5).

A two-receptor model for colicin E1 (ColE1) translocation across the outer membrane of E. coli has been proposed (Masi et al., 2007). ColE1 initially binds to the vitamin B₁₂ receptor BtuB and then translocates through the TolC channel-tunnel, presumably in a mostly unfolded state. In the early events in the import of ColE1, cleavage of colicin requires the presence of the receptor BtuB and the protease OmpT, but not that of TolC. Strains expressing OmpT cleaved ColE1 at K84 and K95 in the N-terminal translocation domain, leading to the removal of the TolQA box, which is essential for ColE1's cytotoxicity. Thus, OmpT degrades colicin at the cell surface to protect sensitive E. coli cells. Secondary binding of ColE1 to TolC depends on primary binding to BtuB, and alterations to residues in the TolC channel can interfere with the translocation of ColE1 but not binding of ColE1 to TolC (Masi et al., 2007).

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