Encyclopedia of Biophysics

Living Edition
| Editors: Gordon Roberts, Anthony Watts, European Biophysical Societies

Bacterial Lipopolysaccharide, OPS, and Lipid A

  • Antonio MolinaroEmail author
  • Cristina De Castro
  • Michelangelo Parrilli
Living reference work entry
DOI: https://doi.org/10.1007/978-3-642-35943-9_78-1

Lipopolysaccharides are amphiphilic molecules indispensable for viability and survival of Gram-negative bacteria, as they heavily contribute to the structural integrity of their outer membrane (OM) and to the protection of the bacterial cell envelope (Di Lorenzo et al. 2015). The highly ordered structure and low fluidity of the LPS layer, stabilized by electrostatic interactions between divalent cations (as Ca2+ and Mg2+) and negatively charged groups present on LPS molecules, are responsible for the increase of permeability to hydrophobic compounds and to higher molecular weight hydrophilic compounds but also for Gram-negative superior resistance to external stress factors. Indeed, only certain antibiotics directed against Gram-negative bacteria, such as polymyxin B, are able to destabilize abovementioned ionic interactions leading to the disruption of membrane integrity. In addition, since they are exposed toward the external environment, LPS molecules participate in crucial mechanisms of host-bacterium interactions as colonization, virulence in the case of pathogen and opportunistic bacteria, adhesion, and symbiosis. Among all these activities, LPS has been shown to be the most potent immunostimulant molecule playing a key role in the pathogenesis of Gram-negative infections triggering the immune system in a wide range of eukaryotic organisms ranging from insects to plants and to humans (De Castro et al. 2012).

LPSs belonging to different bacterial species share a common architecture but possess different chemical structures. LPS is generally built up of three structural and biosynthetically different parts (smooth-type LPS, S-LPS): an O-specific polysaccharide (OPS) covalently linked to an oligosaccharide (core), in turn, linked to a glycolipid moiety (lipid A). OPS might be lacking in some wild-type strains (rough-type LPS, R-LPS, lipooligosaccharide, LOS) (Di Lorenzo et al. 2015) (Fig. 1).
Fig. 1

Sketch of a general structure of LPS

LPS is anchored by its lipid A moiety to the outer membrane, and the carbohydrate part protrudes toward the exterior of the cell. OPS is usually built up of a regular polysaccharide with repeating units (up to 40–50) consisting of two to eight monosaccharides. The lipid A structure consists of a β-(1 → 6)-linked 2-amino-2-deoxy-glucopyranose (GlcN) disaccharide that bears 3-(R)-hydroxy fatty acid residues, as ester-linked at the 3 and 3′ positions and amide-linked at 2 and 2′ positions, which are indicated as primary fatty acid residues (C14:0 3-OH in E. coli) (Fig. 2). The 3-OH, in turn, can be further esterified by secondary fatty acids typically not carrying any other functional group (C14:0 and C12:0 in E. coli). The hydroxyl at position 4′ of the nonreducing GlcNII residue (distal unit) and that of the α-anomeric position of the reducing GlcNI residue (proximal unit) are generally both linked to charged residues, mostly phosphate groups (Molinaro et al. 2015).
Fig. 2

The LPS lipid A structure of E. coli

Both polar head and acyl residue assortments in lipid A may vary in number, type, and distribution and determine the three-dimensional structure, i.e., the conical or cylindrical molecular shape, of lipid A which is correlated to its biological activity, i.e., the binding and recognition by proteins of the innate immune system of both animals and plants (Molinaro et al. 2015; Silipo et al. 2010).

Core oligosaccharide is a complex component of the LPS molecule since it can be characterized by up to 15 monosaccharides which can be organized giving either a linear or a branched structure. The inner part of core region is directly linked to the lipid A, is well conserved, and consists of characteristic monosaccharide residues such as heptoses (L-glycero-D-manno-heptose and D-glycero-D-manno-heptose) and Kdo (3-deoxy-D-manno-octulosonic acid); this latter is considered a diagnostic marker for all Gram-negative bacteria and covalently connects the core oligosaccharide to lipid A backbone with a α-configured ketosidic linkage in almost every LPS investigated to date.

The OPS is the most variable portion of the LPS also within bacteria belonging to the same genus, and for most Gram-negative bacteria, it consists of up to 50 identical repeating oligosaccharide units consisting of two to eight different glycosyl residues (heteroglycans) or of identical monosaccharides (homoglycans). A bacterial strain produces LPSs with O-chains characterized by a wide range of lengths; this different degree of polymerization is responsible for the ladderlike pattern, showed by SDS-PAGE, typical of a S-LPS. The high structural variability of the O-polysaccharide is ascribable to the large number of sugar residues, to their different arrangements (in pyranose or furanose rings, anomeric and absolute configurations) that can build up the repeating units, as well as to the glycosidic sequence and to the presence of noncarbohydrate substituents such as phosphate, amino acids, sulfate, and acyl groups, often present in a nonstoichiometric fashion.

The determination of the primary structure of a LPS is carried out separately on lipid A and OPS (De Castro et al. 2010), chemically split in advance, by means of chemical analyses and biophysical methodologies (gas liquid chromatography coupled with mass spectrometry, GLC-MS; mass spectrometry, MS; and nuclear magnetic resonance, NMR) (De Castro et al. 2010). The determination of the primary structure of LPS implies the rigorous ascertainment of the following points (in parenthesis is also indicated the approach of choice):
  1. (a)

    Quali-/quantitative composition of monosaccharides and fatty acids (GLC-MS)

  2. (b)

    Determination of size of the rings of each sugar residue (GLC-MS)

  3. (c)

    Determination of attachment points of each sugar residue (GLC-MS, NMR)

  4. (d)

    Determination of the absolute configuration of each sugar residue (GLC-MS)

  5. (e)

    Determination of the anomeric configuration of each sugar residue (NMR)

  6. (f)

    Determination of the sequence of monosaccharides in the chain (MS, NMR)

  7. (g)

    Determination of primary and secondary fatty acid location (MS)

  8. (h)

    Determination of nature and location of noncarbohydrate/fatty acid components (MS, NMR, GLC)



  1. De Castro C, Parrilli M, Holst O, Molinaro A (2010) Microbe-associated molecular patterns in innate immunity: extraction and chemical analysis of gram-negative bacterial lipopolysaccharides. Methods Enzymol 480:89–115CrossRefPubMedGoogle Scholar
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  3. Di Lorenzo F, Silipo A, Lanzetta R, Parrilli M, Molinaro A (2015) Bacterial lipopolysaccharides: an overview of their structure, biosynthesis and immunological activity. In: Cipolla L (ed) Carbohydrates chemistry: state-of-the art and challenges for drug development. Imperial College Press, London, pp 57–89Google Scholar
  4. Molinaro A, Holst O, Di Lorenzo F, Callaghan M, Nurisso A, D’Errico G, Zamyatina A, Peri F, Berisio R, Jerala R, Jiménez-Barbero J, Silipo A, Martín-Santamaría S (2015) Chemistry of lipid A: at the heart of innate immunity. Chem Eur J 21:500–519CrossRefPubMedGoogle Scholar
  5. Silipo A, Erbs G, Shinya T, Dow JM, Parrilli M, Lanzetta R, Shibuya N, Newman M-A, Molinaro A (2010) Glyco-conjugates as elicitors or suppressors of plant innate immunity. Glycobiology 20:406–419CrossRefPubMedGoogle Scholar

Copyright information

© European Biophysical Societies' Association (EBSA) 2018

Authors and Affiliations

  • Antonio Molinaro
    • 1
    Email author
  • Cristina De Castro
    • 2
  • Michelangelo Parrilli
    • 1
  1. 1.Department of Chemical SciencesUniversità di Napoli Federico IINaplesItaly
  2. 2.Department of Agricultural SciencesUniversità di Napoli Federico IINaplesItaly

Section editors and affiliations

  • Elizabeth Hounsell
    • 1
  1. 1.School of Biological and Chemical SciencesBirkbeck College, University of LondonLondonUK