Molecular architecture and functioning of the outer membrane of Escherichia coli and other gram-negative bacteria

Publisher Summary This chapter attempts to synthesize the current knowledge of the outer membrane, emphasizing the structural and molecular basis of various functions. The chapter explores that all the bacterial cells except those of mycoplasma and L-forms are surrounded by cell wall. It is a common knowledge that a fundamental difference exists in the structure of cell wall between the Gram-positive and Gram-negative prokaryotes, and that in the latter the cell wall contains an outer membrane layer in addition to the underlying peptidoglycan layer. The outer membrane especially that of Escherichia coli and Salmonella pphimurium, has been studied extensively during the last several years, and at present it claims to understand its structure and function properly at the molecular level. The chapter highlights that together with the erythrocyte membrane, the outer membrane is one of the best studied biological membranes.

The Outer Membrane of Gram-negative Bacteria

The envelope of gram-negative bacteria contains three structures that can be visualized in the electron microscope (FIGURE 1 ) and can now also be separated, which permits their analysis: an inner, or cytoplasmic, membrane that contains lipid and protein; a thin peptidoglycan layer; and an outer membrane that contains lipopolysaccharide (LPS), in addition to protein and lipid. Although gram-positive organisms also have a cytoplasmic membrane and peptidoglycan layer, the outer membrane is unique to gram-negative organisms and confers special properties upon them. This paper will summarize the evidence that the outer membrane provides a permeability barrier that makes gramnegative bacteria less permeable than gram-positive bacteria to a large variety of molecules, which includes many drugs and antimetabolites. This evidence is twofold: chemical, because it has been found that ethylenediaminetetraacetate (EDTA) both increases cell permeability and specifically alters the outer membrane by releasing LPS; and genetic, because some mutants altered in LPS structure exhibit permeability changes and others with increased permeability display outer membrane alteration. A tentative model of the structure of the outer membrane and the aspects of this structure that provide the permeability barrier will be presented. The barrier function of the outer membrane first became accessible to study when it was found that EDTA disrupts this function. In 1958, MacGregor and Elliker discovered that mutants of Pseudornonns nerziginosa resistant to quartenary ammonium compounds become sensitive when exposed to EDTA, and these workers correctly concluded that EDTA increased permeability to these compounds; this observation appears to have been ignored, probably because, as will be discussed below, EDTA alone is much more toxic to this organism than to other gram-negative bacteria. Soon thereafter, Repaske found that EDTA permits lysozyme action on several gram-negative bacteria.g Spheroplasts formed by this method are sensitive to actinomycin, a drug that normally cannot affect intact Escherichia coli:', 1 While pursuing this observation in my laboratory, I found that EDTA alone, without lysozyme, conferred subsequent sensitivity to actinomycin upon the cells,5 and I concluded that EDTA permits entry of the drug. This conclusion was verified by the finding that EDTA-treated, but not control, cells, could bind radioactive actinomycin and that other molecules normally excluded could also enter.'; The permeability increase is very general, as is expected for partial breakdown of a passive permeability barrier. Many substances normally excluded can enter many species of gram-negative bacteria after they are treated with EDTA (TABLES 1 & 2 ) . Of special interest to this conference is the wide range of drugs, unrelated either in their structure or their action, that can attack such

The barrier function of the gram-negative envelope

The outer membrane of gram-negative bacteria such as Escherichia coli serves as a protective barrier that controls the influx and efflux of solutes. This allows the bacteria to inhabit several different, and often hostile, environments. The assembly of the E. coli outer membrane has been difficult to study using traditional genetic and biochemical methods, and how all its components reach the outer membrane after being synthesized in the cytoplasm and cytoplasmic membrane, how they are assembled in an environment that is devoid of an obvious energy source, and how assembly proceeds without disrupting the integrity of this essential cellular structure are all fundamental questions that remain unanswered. Here, we review the new approaches that have led to the recent discovery of components of the machinery involved in the biogenesis of this distinctive cellular organelle.

Advances in understanding bacterial outer-membrane biogenesis

INTRODUCTION STRUCTURAL ORGANIZATION OF THE GRAM-NEGATIVE CELL ENVELOPE ........................................................................ CytovlasmicMembrane. Peptidoglycan-Lipoprotein Complex. Periplasmic Z ne. Outer MembraneLayer. External Layers ........................................................... SYNTHESIS OF THE GRAM-NEGATIVE CELL WALL ....... ................... ASSOCIATION OF ENZYMES WITH SPECIFIC CELL WALL COMPONENTS.. FUNCTION OF CELL WALL-ASSOCIATED ENZYMES ......................... EPILOGUE ....................................... 87 87 89 92 93 96 97 100 101 102

Structure and function of the cell envelope of gram-negative bacteria.

Cell wall and membrane subfractions of the cell envelope of Escherichia coli have been isolated by a procedure involving particle electrophoresis and sucrose gradient density centrifugation. The lipid content of each fraction has been investigated. The individual phospholipids of both fractions are quantitatively similar except that the proportion of lysophosphatidylethanolamine is greater in the wall than in the membrane. Fatty acid analysis of the phospholipids of each fraction revealed that the wall phospholipids contain a greater proportion of palmitic acid. Coenzyme Q is almost exclusively localized in the cell membrane.

W. J. Lennarz

Papers

Distribution of lipids in the wall and cytoplasmic membrane subfractions of the cell envelope of Escherichia coli.