Peptidoglycan types of bacterial cell walls and their taxonomic implications.

Gram-negative bacteria characteristically are surrounded by an additional membrane layer, the outer membrane. Although outer membrane components often play important roles in the interaction of symbiotic or pathogenic bacteria with their host organisms, the major role of this membrane must usually be to serve as a permeability barrier to prevent the entry of noxious compounds and at the same time to allow the influx of nutrient molecules. This review summarizes the development in the field since our previous review (H. Nikaido and M. Vaara, Microbiol. Rev. 49:1-32, 1985) was published. With the discovery of protein channels, structural knowledge enables us to understand in molecular detail how porins, specific channels, TonB-linked receptors, and other proteins function. We are now beginning to see how the export of large proteins occurs across the outer membrane. With our knowledge of the lipopolysaccharide-phospholipid asymmetric bilayer of the outer membrane, we are finally beginning to understand how this bilayer can retard the entry of lipophilic compounds, owing to our increasing knowledge about the chemistry of lipopolysaccharide from diverse organisms and the way in which lipopolysaccharide structure is modified by environmental conditions.

Molecular Basis of Bacterial Outer Membrane Permeability Revisited

Molecular basis of bacterial outer membrane permeability.

Endotoxins of Gram-negative microbes fulfill as components of the outer membrane a vital function for bacterial viability and, if set free, induce in mammalians potent pathophysiological effects. Chemically, they are lipopolysaccharides (LPS) consisting of an O-specific chain, a core oligosaccharide, and a lipid component, termed lipid A. The latter determines the endotoxic activities and, together with the core constituent Kdo, essential functions for bacteria. The primary structure of lipid A of various bacterial origin has been elucidated and lipid A of Escherichia coli has been chemically synthesized. The biological analysis of synthetic lipid A partial structures proved that the expression of endotoxic activity depends on a unique primary structure and a peculiar endotoxic conformation. The biological lipid A effects are mediated by macrophage-derived bioactive peptides such as tumor necrosis factor alpha (TNF). Macrophages possess LPS receptors, and the lipid A regions involved in specific binding a...

Bacterial endotoxin: molecular relationships of structure to activity and function.

Taxonomy relies on three key elements: characterization, classification and nomenclature. All three elements are dynamic fields, but each step depends on the one which precedes it. Thus, the nomenclature of a group of organisms depends on the way they are classified, and the classification (among other elements) depends on the information gathered as a result of characterization. While nomenclature is governed by the Bacteriological Code, the classification and characterization of prokaryotes is an area that is not formally regulated and one in which numerous changes have taken place in the last 50 years. The purpose of the present article is to outline the key elements in the way that prokaryotes are characterized, with a view to providing an overview of some of the pitfalls commonly encountered in taxonomic papers.

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Notes on the characterization of prokaryote strains for taxonomic purposes.

The cDNAs encoding the human bone morphogenetic proteins BMP-2 and BMP-4 in an eukaryotic expression vector were permanently transferred into the murine mesenchymal progenitor cell line C3H10T½. Originally, these cells are known to differentiate into myotubes, adipocytes, and chondrocytes upon the addition of azacytidine. Permanent transfection of genes encoding human BMP-2 and BMP-4 induces differentiation into the osteogenic lineage. The osteogenic differentiation potential of C3H10T½ cells is substantiated by histochemical and genetic analyses of marker genes typical or specific for osteogenesis, including the parathyroid hormone receptor, alkaline phosphatase, osteopontin, osteonectin, and osteocalcin. In addition to osteoblast formation, development into adipocytes and chondrocytes is also observed, suggesting that BMP-2 and BMP-4 induce differentiation into three mesenchymal lineages.

Expression of Human Bone Morphogenetic Proteins-2 or -4 in Murine Mesenchymal Progenitor C3H10T½ Cells Induces Differentiation into Distinct Mesenchymal Cell Lineages

Enterobacterial common antigen (ECA) is a family-specific surface antigen shared by all members of the Enterobacteriaceae and is restricted to this family. It is found in freshly isolated wild-type strains as well as in laboratory strains like Escherichia coli K-12. The family specificity of ECA can be used for taxonomic and diagnostic purposes. ECA is located in the outer leaflet of the outer membrane. It is a glycophospholipid built up by an aminosugar heteropolymer linked to an l-glycerophosphatidyl residue. In a few rough mutants, in addition, the sugar chain can be bound to the complete lipopolysaccharide (LPS) core. Recently, for Shigella sonnei a lipid-free cyclic form of ECA was reported. The genetical determination of ECA is closely related to that of lipopolysaccharide. For biosynthesis of ECA and LPS partly the same sugar precursors and the same carrier lipid is used.

ECA, the enterobacterial common antigen.

The three-dimensional supramolecular structures and the states of order of the acyl chains of lipid A from different Gram-negative species were investigated at 40°C, high water content (80–90%), and different [lipid A]/[Mg2+] molar ratios using synchrotron radiation X-ray diffraction and Fourier-transform infrared spectroscopy. Measurements were made on free lipid A from Salmonella minnesota R595, mono- and bi-phosphoryl, as well as those from the non-enterobacterial strains Rhodobacter capsulatus 37 b4, Rhodopseudomonas viridis F, and Rhodocyclus gelatinosus 29/1. Parallel to differences in their chemical primary structure, the structural polymorphisms and states of order at 37°C of the non-enterobacterial lipid A were found to be different from those of enterobacterial lipid A. A clear correlation between the supramolecular structure and previously determined biological activities was found. Lipid A with a strong preference for lamellar structures (Rb. capsulatus and Rp. viridis) are endotoxically inactive and lack cytokine-inducing capacity; the compounds assuming a mixed lamellar/nonlamellar structure (monophosphoryl lipid A from S. minnesota) are of lower toxicity in vivo, but may induce cytokines in vitro; those lipid A with a strong tendency to form non-lamellar inverted structures (lipid A from S. minnesota and Rc. gelatinosus) exhibit full endotoxicity in vitro and in vivo. In contrast, anti-complementary activity is most pronounced for compounds with lamellar and least expressed for those with inverted structures. The states of order at 37°C vary non-systematically, exhibiting the highest values for lipid A of S. minnesota and the lowest for that of Rc. gelatinosus.



We propose to extend the term ‘endotoxic conformation’, which is used to describe the conformation of a single lipid A molecule required for optimal triggering of biological effects, to ‘endotoxic supramolecular conformation’ which denotes the particular organization of lipid A aggregates in physiological fluids causing biological activity.

Influence of the supramolecular structure of free lipid A on its biological activity

1
R forms of Escherichia coli 0100:K(B) and 0111:K58 were obtained by genetic recombination transferring into the respective S forms a lesion in the his-linked som locus. Such mutants are not able to synthesize O-specific polysaccharide side chains but do have a complete core region.

2
R forms of Citrobacter 010,9b and Arizona 09a,9c were isolated at random as rough looking colonies and characterized as possessing complete core lipopolysaccharide.

3
These R forms and their respective lipopolysaccharides were compared in phage sensitivity, serological properties and in chemical composition with known prototypes of Salmonella and E. coli R mutants (coli R1 and R2) having a complete core structure.

4
According to the phage pattern, the R mutants could be subdivided into distinct groups. One group comprising the R forms of E. coli 0111 and Citrobacter showed a peculiar phage pattern hitherto not observed with other enterobacterial R mutants. The phage pattern of R mutants of Salmonella and Arizona proved to be identical. The R form of E. coli 0100 on the other hand gave the same phage pattern as described for E. coli R2 type mutants.

5
The results of phage typing were fully corroborated by the serological and chemical investigations upon the lipopolysaccharide. In serological studies lipopolysaccharides of all R types except coli R1 were found to cross react with each other, thus indicating structural similarities. In all cross reacting lipopolysaccharide preparations the same sugar constituents namely glucose, galactose, N-acetylglucosamine, l-glycero-d-manno-heptose and 3-deoxy-d-manno-octulosonic acid were present; but quantitative analysis revealed remarkable differences. Striking is the very low content of galactose in the lipopolysaccharide of the R mutants of E. coli 0111 and Citrobacter.

6
The results described show that the E. coli 0111 core represents a further core type which will be designated coli R3.

7
The serological and chemical investigation of the lipopolysaccharide of the Arizona R mutant confirms its identity with the Salmonella core type, thus indicating the close relationship of these bacterial groups.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1970.tb00297.x

Immunochemical studies on core lipopolysaccharides of Enterobacteriaceae of different genera.

Publisher Summary This chapter discusses the analysis of lipopolysaccharides (LPS) of gram-negative bacteria. LPS are characteristic components of the cell wall of all Gram-negative bacteria and of at least some cyanobacteria. They are localized in the outer layer of the outer membrane, and are, in noncapsulated strains, exposed on the cell surface. They contribute to the integrity of the outer membrane and, as has been shown for enteric bacteria, protect the cell against the action of bile salts and (lipophilic) antibiotics. Isolated lipopolysaccharide, upon injection into higher organisms, exhibits a variety of biological (“endotoxic”) activities, such as the induction of fever, changes in the white blood cell count, diarrhoea, or even shock and death. Since the toxic activities are because of substances released from disintegrating bacteria, and not to an excreted exotoxin, lipopolysaccharide has been termed an endotoxin. Thus, the terms lipopolysaccharide, O-antigen, and endotoxin are used synonymously and describe either the chemical structure or discrete biological functions of the same substance.

Hubert Mayer

Papers

Expression of Human Bone Morphogenetic Proteins-2 or -4 in Murine Mesenchymal Progenitor C3H10T½ Cells Induces Differentiation into Distinct Mesenchymal Cell Lineages

ECA, the enterobacterial common antigen.

Influence of the supramolecular structure of free lipid A on its biological activity

Immunochemical studies on core lipopolysaccharides of Enterobacteriaceae of different genera.

6 Analysis of Lipopolysaccharides of Gram-Negative Bacteria