Question

how many outer layers does e. coli (a prokaryote) have ? what are they?

Answer 1

Starting from the outside and proceeding inward the first layer encountered is the OM. The OM is a distinguishing feature of Gram-negative bacteria; Gram-positive bacteria lack this organelle. Like other biological membranes, the OM is a lipid bilayer, but importantly, it is not a phospholipid bilayer. The OM does contain phospholipids; they are confined to the inner leaflet of this membrane. The outer leaflet of the OM is composed of glycolipids, principally lipopolysaccharide (LPS) (Kamio and Nikaido 1976). LPS is an infamous molecule because it is responsible for the endotoxic shock associated with the septicemia caused by Gram-negative organisms (Raetz and Whitfield 2002). The human innate immune system is sensitized to this molecule because it is a sure indicator of infection.

With few exceptions, the proteins of the OM can be divided into two classes, lipoproteins and β-barrel proteins. Lipoproteins contain lipid moieties that are attached to an amino-terminal cysteine residue (Sankaran and Wu 1994). It is generally thought that these lipid moieties embed lipoproteins in the inner leaflet of the OM. In other words, these proteins are not thought to be transmembrane proteins. There are about 100 OM lipoproteins in E.coli, and the functions of most of these are not known (Miyadai et al. 2004; but see below). Nearly all of the integral, transmembrane proteins of the outer membrane assume a β-barrel conformation. These proteins are β sheets that are wrapped into cylinders, and we will refer to these outer membrane proteins as OMPs. Not surprisingly, some of these OMPs, such as the porins, OmpF, and OmpC, function to allow the passive diffusion of small molecules such as mono- and disaccharides and amino acids across the OM. These porins have 16 transmembrane β strands, they exist as trimers (Cowan et al. 1992), and they are very abundant; together they are present at approximately 250,000 copies per cell. Other OMPs, such as LamB (18 transmembrane β strands) (Schirmer et al. 1995) or PhoE (16 transmembrane β strainds) (Cowan et al. 1992), exist as trimers as well and they function in the diffusion of specific small molecules, maltose or maltodextrins and anions such as phosphate respectively, across the OM. When induced by the presence of maltose or phosphate starvation, respectively, these proteins are very abundant as well. OmpA is another abundant OMP. It is monomeric, and it is unusual in that it can exist in two different conformations (Arora et al. 2000). A minor form of the protein, with an unknown number of transmembrane strands, can function as a porin, but the major, nonporin form has only eight transmembrane strands, and the periplasmic domain of this form performs a largely structural role (see later discussion). An additional class of OMPs, which are larger β-barrels (20–24 transmembrane β strands), but are present at much lower levels, function as gated channels in the high affinity transport of large ligands such as Fe-chelates or vitamins such as vitamin B-12 (for review see Nikaido 2003).

The OM is essential for the survival of E. coli, but it contains only a few enzymes. For example, there is a phospholipase (PldA) (Snijder et al. 1999), a protease (OmpT) (Vandeputte-Rutten et al. 2001), and an enzyme that modifies LPS (PagP) (Hwang et al. 2002). The active site of all of these enzymes is located in the outer leaflet, or it faces the exterior of the cell (OmpT). Mutants lacking any of these enzymes exhibit no striking phenotypes. The only known function of the OM is to serve as a protective barrier, and it is not immediately obvious why this organelle is essential. But what a barrier it is. Salmonella, another enteric bacterium, can live at the site of bile salt production in the gall bladder (Sinnott and Teall, 1987), and it is generally true that Gram-negative bacteria are more resistant to antibiotics than are their Gram-positive cousins. Indeed, some Gram-negative bacteria, such as Pseudomonas, are notorious in this regard.

LPS plays a critical role in the barrier function of the OM. It is a glucosamine disaccharide with six or seven acyl chains, a polysaccharide core, and an extended polysaccharide chain that is called the O-antigen (Raetz and Whitfield 2002). Traditionally pathogenic E. coli are classified by the antigenic properties of their O-antigen and the major protein (flagellin, termed H) component of the flagella (see later discussion). Hence, E. coli O157:H7. LPS molecules bind each other avidly, especially if cations like Mg++ are present to neutralize the negative charge of phosphate groups present on the molecule. The acyl chains are largely saturated, and this facilitates tight packing. The nonfluid continuum formed by the LPS molecules is a very effective barrier for hydrophobic molecules. This coupled with the fact that the porins limit diffusion of hydrophilic molecules larger than about 700 Daltons, make the OM a very effective yet, selective permeability barrier (Nikaido 2003).