Polysaccharides, Microbial G Morris and S Harding, University of Nottingham, Sutton Bonington, UK
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Polysaccharides, Microbial G Morris and S Harding, University of Nottingham, Sutton Bonington, UK ª 2009 Elsevier Inc. All rights reserved. Defining Statement Dextrans Introduction Scleroglucan and Schizophyllan Xanthan Bacterial Alginates Xylinan (Acetan) Bacterial Hyaluronic Acid Gellan, XM6, and Curdlan Polysaccharide Vaccines Cyanobacterial Polysaccharides Conclusion Pullulan Further Reading Glossary molecular weight averaged over the number capsular polysaccharide (CPS) An exocellular concentrations of each species. It can be determined by polysaccharide secreted in such a large amount that it techniques such as osmotic pressure, size exclusion encapsulates a microorganism, facilitating adhesion chromatography coupled to multiangle light scattering and helping screen from immune response. Sometimes (SEC-MALLs) or analytical ultracentrifugation. a distinction is made with more loosely attached molecular weight – weight average, Mw (Da or À1 polysaccharides ‘released polysaccharide’ (RPS). g mol ) In a mixture of macromolecules Mw is the epimerase An enzyme that converts mannuronic acid molecular weight averaged over the weight residues to guluronic acid residues in alginates. concentrations of each species. It can be determined by intrinsic viscosity, [h] (ml gÀ1) A measure of a techniques such as size exclusion chromatography particle’s conformation and volume occupancy in coupled to multiangle light scattering (SEC-MALLs) or solution. analytical ultracentrifugation. Mark–Houwink ‘a’ parameter A measure of the non-Newtonian fluid A fluid in which viscosity conformation of a macromolecule, and determined from changes with applied strain, an example of which is measurements of solution viscosity and molecular shear thinning. weight. Limits are 0 (sphere), 1.8 (rod), with values persistence length, Lp (nm) A measure of the flexibility 0.5–0.8 for a flexible coil-like molecule. of a linear macromolecule ranging from 2 nm for a very molecular weight (Da or g molÀ1) Popular terminology flexible molecule to 200 nm for very stiff. for ‘molar mass’ – the mass of 1 mol (6.023 Â 1023) polydispersity index The ratio of the weight average molecules. molecular weight to the number average molecular molecular weight – number average, Mn (Da or weight, Mw/Mn. Minimum value ¼ 1. À1 g mol ) In a mixture of macromolecules Mn is the Abbreviations GRAS generally regarded as safe CPS capsular polysaccharide HA hyaluronic acid GDP guanosine diphosphate RPS released polysaccharide Defining Statement properties. We consider the structure, properties, extraction, production, modification, and applications of commercially Microbial polysaccharides are used for food, pharmaceutical, available microbial polysaccharides including xanthan, xyli- and medical applications: this wide range of usefulness nan, gellan, curdlan, pullulan, dextran, scleroglucan, derives from the great diversity in structural and functional schizophyllan, and cyanobacterial polysaccharides. 482 Applied Microbiology: Industrial | Polysaccharides, Microbial 483 Introduction They may also prevent phagocytosis by other micro- organisms or the cells of the immune system. The capsular Over the last two decades there has been an expanding polysaccharides (CPSs) are often highly immunogenic, and interest in polysaccharides produced extracellularly by may have evolved their unusual diversity as a way of microorganisms for food, pharmaceutical, and medical avoiding antibody responses: advantage of this feature can use, including vaccines. The wide range of their useful- be taken in the development of vaccines. They also have a ness derives from the great diversity in structural and role in adhesion and penetration of the host. In the case of functional properties even though they are built up from plants this involves interaction with polysaccharide struc- very similar building blocks: the pyranose or furanose tures in the cell walls and there are possibilities of specific carbohydrate ring structure. One can have rod-shaped interactions. Plant lectins (glycoproteins) that have specific molecules like schizophyllan and scleroglucan from the binding properties with respect to carbohydrate structures fungi Schizophyllan commue and Sclerotium rolfsii, respec- may play a part in this and in the general defense of plants tively, linear random coil type structures like pullulan against bacterial infection. from the fungus Aureobasidium pullulans, and intermediate Some secreted polysaccharides can be involved in structures such as alginates from the bacterium pathogenicity. Pseudomonas aeruginosa, commonly found in Azotobacter vinelandii. One can have highly electronegative respiratory tract infections, produces alginate which con- or ‘polyanionic’ molecules like xanthan secreted by tributes to blockage in the respiratory tract, and leads to Xanthomonas campestris or neutral polysaccharides like further infection, and similar blockages of phloem in plants dextrans from the bacteria Leuconostoc mesenteroides. have also been described. However, the secreted polysac- Polysaccharides made by microrganisms are secreted charides themselves present virtually no known toxicity from the cell to form a layer over the surface of the problems and many are harvestable at low cost in large organism, often of substantial depth in comparison with quantities, making them attractive for biotechnological use. the cell dimensions (Figure 1). Because of their position Microbial products are very important in all aspects of they are characterized as exopolysaccharides, to distin- polysaccharide biotechnology. They range from bulk guish them from any polysaccharides that might be found materials such as xanthan, priced at about US$14 per kg, within the cell. The functions are thought to be mainly used mainly in food applications, to cyclic dextrans, protective, either as a general physical barrier preventing valued at about US$50 per kg and used in high-value access of harmful substances, or more specific as a way of applications in research and pharmaceuticals. binding and neutralizing bacteriophage. In appropriate Certain microbes are known to produce nearly all the environments they may prevent dehydration. major plant polysaccharides such as glucans, alginate-like materials, and even cellulose – as well as the complex bacteria-specific materials. Genetic manipulation of bac- teria has been studied for longer and is in general much easier than for higher organisms, so that they are an P obvious target for both manipulation of biosynthetic path- ways and the expression of heterologous genes to produce especially desirable enzymes. Polysaccharides are not of course under the direct control of genetic material in the way that enzymes are, and must be approached indirectly by manipulating the biosynthetic or degradation path- ways by way of the enzymes responsible: there is currently great scope for this approach. In addition, since there are now fermentation processes involving the large-scale growth of bacteria, all the advances in this field can be achieved by biotechnology. There are also limitations. It is unlikely that large- scale fermentation could ever compete with existing pro- cesses for pectin manufacture, for example, since Figure 1 Exocellular or ‘capsular’ polysaccharide layer processors currently use waste by-products from other (labeled P) from Streptococcus pneumoniae. Adapted from processes as their raw material. This is quite apart from Skov-Sørensen UB, Blom J, Birch-Andersen A, and Henrichsen J the inherent costs of large-scale fermenters in comparison (1988) Ultrastructural localization of capsules, cell wall with simple extraction processes. It follows that bacterial polysaccharide, cell wall proteins and F antigen in pneumococci. Infection and Immunity 56: 1890–1896. Reproduced with products must have particularly useful properties in order permission from the American Society for Microbiology. to justify their probable greater cost. 484 Applied Microbiology: Industrial | Polysaccharides, Microbial This article looks at the production, properties, and trisaccharide involving mannose and acetylated biomodifications where appropriate of a selected range mannose: of important microbial polysaccharides because it is impossible to cover each substance comprehensively. -D-Manpð1!4Þ -D-GlcpA Instead, we focus on xanthan as our main example ð1!2Þ -D-Manp-6-OAcetylð1!3Þ because of its commercial importance but also consider with some of the terminal -D-Manp residues substituted xanthan-like exopolysaccharides such as gellan, xm6, – at positions C4 and C6 by pyruvate (CH3.CO.COO ). curdlan, xylinan, and the increasingly important group Thus xanthan (except under highly acidic conditions) is, of cyanobacterial polysaccharides – also xanthan-like – like alginate, a highly charged polyanion. Figure 2(a) as well as pullulan, dextrans, scleroglucan, schizophyllan, shows the Haworth projection of a (pyruvalated) alternate microbial hyaluronic acid (HA), and bacterial alginates repeat section of xanthan. as representative examples. We complete our review It is not only charge repulsion effects that convey on with a look at the use and potential further use of xanthan an extended rigid rod conformation. X-ray fiber exopolysaccharides as vaccines against serious diseases diffraction studies have shown the presence of some helical such as meningitis in infants. structure although there is still uncertainty as to whether this helix is a coaxial duplex or just a single chain one, with the projecting trisaccharides