Alginate Production: Precursor Biosynthesis, Polymerization and Secretion

Rehm, Bernd H. A.

doi:10.1007/978-3-540-92679-5_2

Cited by 59 publications

(63 citation statements)

References 69 publications

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“…Furthermore, the prolonged time spent in stationary phase for the 48-h treatments, where resource competition is much greater, might increase metabolic costs associated with this particular phenotype. As exopolysaccharide and alginate production is metabolically expensive (Linton, 1991;Rehm, 2009), and resources are increasingly depleted through time, the resultant competition between genotypes is greater, resulting in a less favourable environment for the mucoid phenotype. The three other phenotypes that are routinely recovered from this experimental system, namely SM, WS and FS, were also recovered here from treatments with and without phage (Rainey and Travisano, 1998;Rainey, 2002a, 2002b), suggesting that the mucoid phenotype of P. fluorescens is the only phenotype exclusively associated with phage predation in this experimental context and is not a general adaption to other experimental variables.…”

Section: Discussionmentioning

confidence: 99%

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25

2011

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The effects of co-evolution with lytic phage on bacterial virulence-related traits are largely unknown. In this study we investigate the incidence of the mucoid phenotype of the bacterium Pseudomonas fluorescens SBW25 in response to co-evolution with the lytic phage phi2 (/2). The mucoid phenotype of Pseudomonas spp. is due to overproduction of alginate and is a considerable virulence factor contributing to the intractability of infections most notably in cystic fibrosis (CF) lung, but also in pathogenic infections of plants. Our data show that this phenotype can evolve as an adaptive response to phage predation and is favoured under specific abiotic conditions, in particular a homogenous spatial structure and a high rate of nutrient replacement. The mucoid phenotype remains partially sensitive to phage infection, which facilitates 'apparent competition' with phage-sensitive competitors, partially offsetting the costs of alginate production. Although P. fluorescens SBW25 is not a pathogen, several key characteristics typical of Pseudomonas aeruginosa clinical isolates from CF lung were noted, including loss of motility on mucoid conversion and a high rate of spontaneous reversion to the wild-type phenotype. Although the genetic mechanisms of this phenotype remain unknown, they do not include mutations at many of the commonly reported loci implicated in mucoid conversion, including mucA and algU. These data not only further our understanding of the potential role phage have in the ecology and evolution of bacteria virulence in both natural and clinical settings, but also highlight the need to consider both biotic and abiotic variables if bacteriophages are to be used therapeutically.

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Section: Discussionmentioning

confidence: 99%

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25

2011

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“…Alginate has a variable molecular weight, depending on the enzymatic control during its production and the degree of depolymerization caused by its extraction. Typically, commercial alginates have an average molecular weight of approximately 200,000 Da, but alginates with values as high as 400,000-500,000 Da are also available (Rehm, 2009). The physico-chemical properties of alginate have been found to be highly affected by the M/G ratio as well as by the structure of the alternating zones, which can be controlled by enzymatic pathways (Coviello et al, 2007).…”

Section: Alginatementioning

confidence: 99%

“…Recently, the combination of different epimerases has been used as a fundamental tool in order to create specific engineered alginates with any desired block length and composition (Rehm, 2009). Moreover, alginate has a large number of free hydroxyl and carboxyl groups distributed along the backbone, which are highly reactive and turn it into an ideal candidate for being appropriately modified by chemical functionalization.…”

Section: Alginatementioning

confidence: 99%

Polysaccharide-Based Nanoparticles for Controlled Release Formulations

Martı́nez¹,

Fernandez²,

Pérez³

et al. 2012

The Delivery of Nanoparticles

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“…C-di-GMP stimulates the biosynthesis of several extracellular polysaccharides important for biofilm formation, including alginate and PNAG (Itoh et al, 2008;Rehm, 2009;Steiner et al, 2013). While the mechanism for activating PNAG biosynthesis most likely differs from BcsA (Steiner et al, 2013), alginate and cellulose synthases share a strikingly similar organization (Keiski et al, 2010).…”

Section: Discussionmentioning

confidence: 99%

“…A processive mechanism requires that the elongated polymer is translocated after each glycosyl transfer, such that the polymer's newly added sugar unit becomes the acceptor in a subsequent reaction. Because all known processive GTs are TM channel-forming enzymes (Merzendorfer, 2006;Rehm, 2009;Hubbard et al, 2012), the translocation of the polymer into the TM channel between catalytic steps also gives rise to secretion.…”

Section: Introductionmentioning

confidence: 99%

Structural Studies of Biopolymer Membrane Transport

Morgan¹

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The selective transport of specific substrates across the membrane is an essential part of biology.Organisms have evolved a basic mechanism for the transport of ions and small molecule such as sugars across the membrane. This mechanism is termed 'alternating access' and involves the alternating exposure of a substrate-binding site to either side of the membrane. While the molecular details of 'alternating access' have been revealed for a number of different transporters, this scheme seems infeasible for the transport of long biopolymers such as nucleic acids, proteins, and polysaccharides because 'alternating access' requires that the transporter forms a binding site that is large enough to bind the entire substrate. Here, I present novel data addressing the transport mechanism for two bacterial biopolymer transporter proteins: Bacterial Cellulose Synthase which synthesizes and secretes the polysaccharide, cellulose; and PrtD, an ABC transporter which is involved in the secretion of an extracellular protease.Cellulose is a linear polymer of glucose units that forms a major component of plant cell walls as well as many bacterial biofilms. In bacteria, the components required for cellulose biosynthesis are encoded in a single operon minimally made up of the genes bcsA, bcsB, bcsC, and bcsZ. BcsA is an integral inner-membrane (IM) glycosyltransferase enzyme that is responsible for coupling the synthesis of cellulose with its transport across the IM. BcsB is a periplasmic protein with Cterminal IM anchor, and BcsB forms a complex with BcsA (BcsA-B) that is sufficient for in vitro cellulose synthesis. BcsZ is a periplasmic cellulase enzyme, and BcsC is an outer-membrane β-barrel that presumably forms a pore for the cellulose polymer to cross the outer membrane.iii BcsA-B activity is stimulated by the bacterial signaling molecule, cyclic-di-GMP (c-di-GMP), which is a key regulator of biofilm formation. I present crystal structures of the c-di-GMP-bound BcsA-B complex in the presence and absence of UDP, a competitive inhibitor and substrate mimic. The structures reveal that c-di-GMP releases an auto-inhibited state of the enzyme. A salt bridge stabilizes one of the signature c-di-GMP-binding Arg residues in a position to tether a conserved 'gating loop' in front of the active site. The binding of c-di-GMP releases the tether and allows substrate to access the active site. Additionally, the UDP-bound structure reveals an additional role for the 'gating loop' in coordinating substrate at the active site. Functional experiments confirm the structural interpretation by revealing that disruption of the auto-inhibitory salt bridge by mutagenesis generates a constitutively-active cellulose synthase. The mechanistic insights presented here represent the first examples of how c-di-GMP allosterically modulates enzymatic functions.To address the mechanism by which BcsA-B transports cellulose across the IM, I use in crystallo enzymology with the c-di-GMP-bound BcsA-B crystals. Because crystallized BcsA-B is catalytically a...

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Alginate Production: Precursor Biosynthesis, Polymerization and Secretion

Cited by 59 publications

References 69 publications

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25

Polysaccharide-Based Nanoparticles for Controlled Release Formulations

Structural Studies of Biopolymer Membrane Transport

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